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PTC-0200 DNA Engine & PTC

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1. Alpha unit or base needs ser vicing Contact MJ RESEARCH or your local distributor 11 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual Error Message HS Sensor Fault Alt Control Mode HS Sensor Fault Left Ignored HS Sensor Fault Right Ignored Internal Fan Fault Return for Service Lid Sensor Fault Lid Was Disabled L R Power Imbalance Memory Is Corrupt No Memory Available Probe Sensor Fault Used Calc Control 11 4 Cause and Result Alpha unit heat sink sensor is not functioning properly so machine has begun estimating heat sink temperature based on other tem peature sensor readings Same as previous Same as previous Fan inside base is not working so Alpha unit has been shut down Heated lid is giving unexpected temperature readings so base has turned it off Protocols can still be run but reactions may fail be cause of condensation in vessels Thermoelectric units in Alpha unit block are not heating uniformly so machine begins delivering equal power to all heating ele ments Protocols may still be run but reactions may fail due to un even block temperatures Rarely seen message indicating that memory has been corrupted by a static shock or other unusual electronic incident All available memory has been filled Either probe sensor is not working properly or a probe control pro tocol was run without probe being installed Machine has switched temperatu
2. on the circuit board and slide the latch to lock the pins in place e Press both cables firmly into the two strain relief holes on the Dock Connector s front side fig 13 2 e Turn the Remote Dock upside down You will see a male multi pin connector labeled Power Port and a female multi pin connector la beled Data Port fig 13 3 e Connect the multi pin power cable s female end to the connector la beled Power Port and slide the latch to lock the pins in place e Attach the data cable s DOCK DATA end to the connector labeled Data Port and slide the latch to lock the pins in place e Turn both units back over e The Dock Connector mounts in the base in the same manner as a regular Alpha unit see pp 4 4 4 6 Figure 13 4 Fan power supply connected in series E 009 135 MW DNA Engine amp DNA Engine Tetrad Operations Manual Operation 13 6 e Attach the wall plug power cord to the fan power supply e Attach the fan power supply s round jack to either of the round con nectors on the back of the Remote Dock fig 13 4 e The fan power supply produces sufficient current such that up to three additional fans can be daisy chained to the initial Remote Dock via the round jack power cords supplied Attach one end of the cord to the initial unit s free round connector Attach the other end to either of the round conne
3. 4 a 4 KXX a a a a a a a a AT AT A ao AT _ E EA y a Ok A a SO O ee ee F a A L g o A I g o A I g o aro y r _ PTC 0200 DNA Engine amp A PTC 0225 DNA Engine Tetrad Operations Manual go a Version 4 0 f a a La 4 E a O e z Mi id Xu pan gt 4 SLUT G ie j WOT hs A a mM A Oe f a o M z E AXN a Ma P MJ Research a 4 f y a ON A 0543405 EY ha ZE Y Dar E Y Dar ZE Y Dar o pe E oE ae ero a E T y wa T q RQ T q RQ T RRQ a SJ ess a g s a SJ ON S oe PTC 0200 DNA Engine amp PTC 0225 DNA Engine Tetrad Operations Manual Version 4 0 MJ Research Inc Waltham Massachusetts Copyright 1999 MJ Research Inc All rights reserved Reproduction in any form either print or electronic is prohibited without written permission of MJ Research Inc DNA Engine DNA Engine Tetrad Alpha Hot Bonnet Power Bonnet Microseal Chill Out Self Seal and Twin Tower are trademarks of MJ Research Inc 05434 05 Thermal cyclers can be used for a number of purposes including the polymerase chain reaction PCR PCR is covered by patents owned by Hoffmann La Roche Inc and F Hoffmann La Roche Ltd who have granted exclusive and nonexclusive licenses for some types of applications Roche and its licensees provide end user licenses within their respective fields These licenses have dif ferent terms depending on the particular application of PCR and differ
4. Select Gradient Calculator and press the Proceed key 50 00 Lower Temp C 70 02 Upper Temp C Enter lower limit temperature of the gradient using the number keys and then press the Proceed key Enter the upper temperature for the gradient using the number keys and Proceed key You may enter inte gers without decimal points The following screen will be displayed Gradient Calculator Column 1 50 0 left Column 2 50 59 Column 3 51 5 Column 4 53 2 Column 5 55 52 Column 6 58 4 Column 7 61 8 Column 8 64 6 Column 9 66 82 Column 10 68 4 Column 11 69 62 Column 12 70 0 right You may scroll up and down the screen using the Select keys When scrolling the Gradient Calculator line will be held constant while the lower 3 lines can be manipulated up or down 6 11 MW DNA Engine amp DNA Engine Tetrad Operations Manual 6 12 Entering a Ramp Step To enter a ramp step select Ramp from the Enter Menu The first Ramp screen will be displayed Enter CUSTOM1 3 Rate C Ss _ The second line of this screen shows the number of the step being pro grammed 3 is used in the example above The last line of the screen allows a ramp rate in degrees Celsius per second to be entered for the step Use the keyboard to enter any rate up to 2 5 C sec 1 C sec is used in the example below Enter CUSTOM1 Note If a ramp rate faster than the Alpha unit s maximum rate o
5. ing sensors to monitor block tem perature None of the temperature sensors in Alpha unit are working proper ly so Alpha unit has been shut down Action No action is necessary Protocol resumes running when power is restored Results may or may not be affected depending on whether power failed in an ear ly or a late cycle and whether the power was restored before the sample cooled excessively Determine machine s software version number see p 8 9 If number is below 1 1M contact MJ RESEARCH or your local dis tributor to request a software version upgrade Alpha unit and or base needs servicing Contact MJ RESEARCH or your local distributor Alpha unit needs servicing soon Contact MJ RESEARCH or your local distributor Alpha unit and or base needs servicing Contact MJ RESEARCH or your local distributor Error Message Calc Control Probe Mode Invalid Gradient Fault Service Alpha Soon Gradient Program Invalid Alpha Type Heated Lid Fault Program Terminated HS Overheated Program Terminated HS Overheating Check Air Flow HS PS Sensor Fault Program Terminated Cause and Result Runtime error indicating a eradient step is being progammed under Probe Control mode Displayed at end of run Indicates gradient was not achieved within 20 secs of block reaching target Runtime error indicating gradient is being programmed for an Alpha module other than a 96V Lid sensor failed
6. trated in figure 4 x over 30 seconds can be saved per cycle by using thin walled tubes for an overall savings of 15 minutes in a 30 cycle run Sealing Sample Vessels Steps must be taken to prevent the evaporation of water from reaction mixtures during thermal cycling to avoid changing the concentration of reactants Only a layer of oil or wax will completely prevent evaporation from sample vessels However an adequate degree of protection can be achieved by sealing with Microseal film or caps then cycling the samples using the heated lid to prevent condensation refluxing Sealing with Oil or Wax Mineral oil silicone oil paraffin wax or Chill out liquid wax may be used to seal sample vessels Use only a small amount of oil or wax 1 3 drops 15 50ul are usually sufficient Include this volume in the total volume when setting up a calculated control protocol see Setting Up a Calculated Control Protocol chapter 5 Use the same amount of oil or wax in all sample vessels to ensure a uniform thermal profile Most paraffin waxes solidify at room temperature The wax can then be pierced with a micropipette and the samples drawn off from below the wax Silicone oil and mineral oil can be poured off or aspirated from tubes if the samples are first frozen 15 to 20 C The samples are usually pure enough for analysis without an extraction Chill out liquid wax available from MJ RESEARCH is an easy to use alter native
7. 105 0 C then press Proceed The DNA Engine will incubate the sample at the specified temperature until Cancel or Stop is pressed When the sample block reaches the incubation temperature a timer be gins running in the lower right hand corner of the screen To stop and start the timer press Pause Y Tip The Pause feature is useful if you need to temporarily remove samples that must be incubated for a precise period of time Pausing the timer while samples are not in the block allows you to track the exact duration of their incubation Creating Programs The Elements of a Program 6 2 Designing a New Program 6 2 Translating a Protocol into a Program 6 2 Using the GoTo Step to Write Short Programs 6 3 Choosing a Temperature Control Method 6 3 Calculated Control 6 4 Block Control 6 4 Probe Control 6 4 Modifying Block and Probe Control Programs for Calculated Control 6 5 Modifying a Program Designed for a Different Machine 6 5 Entering a New Program 6 5 Initiating the Program 6 5 Naming the Program 6 6 Choosing a Temperature Control Method 6 6 Entering the Program s Steps 6 6 Entering a Temperature Step 6 7 Entering a Gradient Step 6 9 Editing a Gradient Step 6 10 Reviewing a Gradient Program 6 10 Using the Gradient Calculator 6 10 Factory Installed Gradient Program 6 12 Entering a Ramp Step 6 12 Entering a GoTo Step 6 14 Entering the End Step 6 15 Modifying a Program Step with the Opti
8. The first GoTo screen will be displayed Enter CUSTOM1 5 GoTo GoTo step _ The second line of this screen shows the number of the step being pro grammed 5 is used in the example above The last line of the screen allows entry of the number of the step the program should cycle back to Enter the number of the step the program should cycle back to 1 is used in the example below Enter CUSTOM1 5 GoTo GoTo step 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual 6 14 Press Proceed The step number will move to the second line of the screen and a line allowing an additional number of cycles to be entered will be displayed Enter CUSTOM1 3 GoTo 1 Addtnl cycles Enter the additional number of times the program should cycle back to the step 3 is used in the example below Enter CUSTOM1 3 GoTo 1 Addtnl cycles Press Proceed The number of additional cycles will move to the sec ond line of the screen and a confirmation menu will be displayed on the last line Enter CUSTOM1 3 GoTo 1 3 times OK _YES No Select one of the displayed choices then press Proceed e Yes accepts the step and displays the Enter Menu again Use the En ter Menu to enter the next step in the program e No allows reentry of the step number and number of additional cycles Entering the End Step To enter the End step select End from the Enter Menu The single End screen will be disp
9. adjusting lid pressure 4 8 4 9 temperature control methods 8 9 Increment option See Programs options types of Instant incubation 5 10 L Layout back view 2 3 bottom view 2 3 control panel 2 2 front view 2 2 M Menus Edit Menu 7 3 Enter Menu the 6 6 Files Menu 8 2 Main Menu 4 2 Options Menu 6 11 menus Setup Menu 8 7 Microseal A film 4 8 M sealing mat 4 8 P pads 4 14 N Networking general information 9 2 with computer as controller 9 4 setting up an IEEE 488 network 9 5 setting up an RS 232 network 9 5 with DNA Engine as controller 9 2 9 4 assigning address numbers 9 2 connecting machines 9 2 running protocols 9 3 In 2 O Oil use of in block See Sample vessels Operation turning machine on 4 2 P Packing checklist 3 2 Passwords See Utilities files assigning password to folder Peltier effect C 1 Ports 4 3 TEEE 488 2 3 parallel printer 2 3 RS 232 2 3 Power Bonnet 2 4 Power cord location of jack 2 3 plugging in 3 2 Power supply requirements 3 3 acceptable power cords 3 3 Probe adding representative sample calculating amount of oil to add 4 12 choosing type of oil 4 12 how to add oil 4 12 connecting to block 4 13 customizing probe vessel 4 11 detecting faulty 4 13 function of 4 11 layout 4 11 loading into block 4 13 thermistor 4 11 probe adding representative sample 4 12 silicone oil as representative sample 4 17 Programming Choosing a temperature control method 6 6 delet
10. to facilitate robotic operation as long as the airway beneath the unit is equivalent to the airway the unit would have with the feet attached Usually a hole will need to be cut to allow air to flow to the the cooling fan Figure 13 5 is a template for flush mounting the Remote Dock Power Supply The Alpha unit mounted in each Remote Dock is powered from the PTC 200 or PTC 225 base The Remote Dock s fan is powered externally and a power supply is provided that requires power from 90 250 VAC and 47 to 63 Hz with a grounded outlet Air Supply Alpha units being operated in the remote configuration have no op erating constraints that do not also apply to normal operations in the PTC 200 or PT C 225 Figure 13 3 Attachment of power and data cables 13 4 Dock connector Remote dock The Remote Alpha Dock System M Installation e Turn the Dock Connector upside down so that the green circuit board is visible Check that the ground lead remains attached at both ends fig 13 1 Note the two female multi pin sockets one labeled DATA and the other POWER e Connect the multi pin power cable s male end to the female socket labeled POWER and slide the latch to lock the pins in place e Both of the data cable s multi pin connectors are male one is labeled CONNECTOR DATA and the other DOCK DATA Attach the CONNECTOR DATA end to the female connector labeled DATA
11. 200 DNA Engine or PTC 225 DNA Engine Tetrad Doing so will void your warranties and can put you at risk for electrical shock Return the PTC 200 DNA Engine or PTC 225 DNA Engine Tetrad to the factory US customers or an authorized distributor all other customers if re pairs are needed All Alpha unit blocks can become hot enough during the course of normal op eration to cause burns or cause liquids to boil explosively Wear safety goggles or other eye protection at all times during operation The PTC 200 DNA Engine and PTC 225 DNA Engine Tetrad incorporate neu tral fusing which means that live power may still be available inside the ma chines even when a fuse has blown or been removed Never open the PTC 200 DNA Engine or PTC 225 DNA Engine Tetrad base you could receive a serious electrical shock Opening the base will also void your warranties Never remove an Alpha unit from the PTC 200 DNA Engine or PTC 225 DNA Engine Tetrad with the power turned on and a program running Doing so can cause electrical arcing that can melt the contacts in the connector joining the Alpha unit to the PTC 200 DNA Engine or PTC 225 DNA Engine Tetrad Safe Use Guidelines The PTC 200 DNA Engine and PTC 225 DNA Engine Tetrad are designed to be safe to operate under the following conditions e Indoor use e Altitude up to 4000m A 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual e Ambient temperature 5 31 C e Maximum relative humidit
12. 6 0 5 ml Tubes 4 6 0 2 ml Tubes 4 6 Microplates 4 6 Sealing Sample Vessels 4 7 Sealing with Oil or Wax 4 7 Sealing with the Hot Bonnet and Caps or Film Sheets 4 7 Adjusting the Hot Bonnet s Lid Pressure 4 8 Loading Sample Vessels into the Block 4 9 Using Oil to Thermally Couple Sample Vessels to the Block 4 9 Using the Optional Probe 4 10 Customizing the Probe Vessel 4 10 Adding the Oil 4 11 Loading and Connecting the Probe 4 11 Detecting a Faulty Probe 4 12 Appendix 4 A Tube Microplate and Sealing System Selection Chart 4 13 Appendix 4 B Safety Warning Regarding Use of 45 Nucleotides 4 14 Appendix 4 C Using Silicone Oil in the Probe Tube 4 16 4 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual Turning the DNA Engine On Move the power switch to 1 the On position In most The fan will turn on and the power light on the keyboard will glow red In most cases a self test of the heat pumps will begin running see note below Its progress is tracked on a screen in the LCD window PTC 200 Self testing This screen disappears in 1 minute If a problem is detected the display shows an error message Note If the Alpha unit s block or heat sink is not at ambient tempera ture typically because the Alpha unit was recently in use the machine will skip the self test If the self test does not detect any problems the Main Menu is displayed PTC 200 _RUN Enter List Edit Files
13. A or a B if a dual block is involved For example the designa tion for a single block Alpha unit present in quadrant 1 of machine 17 is 17 1 The designation for block B in a dual block Alpha unit present in quadrant 2 of the same machine is 17 2B Using the Utilities for the DNA Engine Tetrad The DNA Engine Tetrad has the same utilities as the DNA Engine see chapter 8 12 9 MW DNA Engine amp DNA Engine Tetrad Operations Manual Troubleshooting the DNA Engine Tetrad The Tetrad is subject to the same problems with operations protocols and reaction results as the DNA Engine see chapter 11 Maintaining the DNA Engine Tetrad Cleaning Clean the DNA Engine Tetrad according to the instructions in chapter 10 for cleaning the DNA Engine Changing the Fuses Each quadrant of the Tetrad is protected by a fuse mounted in a plug on the back panel of the base fig 12 6 Blown fuses may be replaced by any 15A fast acting fuse 0 25 x 1 25 3AB or 3ABC To replace a fuse first unplug the power supply from the wall Twist the fuse cap a quarter turn to the left and pull out the fuse plug Replace the blown fuse Press the plug back into its socket and twist it a quarter turn to the right to seat it The plug is spring loaded so firm pressure may be required Warning The DNA Engine incorporates neutral fusing which means that live power may still be available inside the unit even when a fuse has blown or been removed
14. Engine s parallel interface is compatible with most serial and laser printers the Epson LX 810 dot matrix printer parallel interface and dot matrix print ers using interfaces similar to the Epson s Use a standard DB 25 Centronics parallel interface cable to connect the DNA Engine s parallel port to the printer s parallel port Anull modem serial cable will usually be required to connect the DNA Engine s serial port to a printer s serial port Use the Setup utilities to choose a printer port for the DNA Engine see Choosing a Printer Port p 8 8 Select List from the Main Menu then press Proceed Locate the program to be printed see Locating a Stored Program p 8 2 then press Pro ceed The screen asking whether the program should be sent to the printer will be displayed see above Select Yes then press Proceed The program will be printed Use these utilities available from the Setup Menu to accomplish the following tasks e Choose a remote port and a printer port e Choose a temperature control method for the Hot Bonnet s heated lid e Look up the DNA Engine s software version number To display the Setup Menu select Setup from the Main Menu then press Proceed Setup _REMOTE Printer Lid Version Choosing a Remote Port To choose either the serial port or the IEEE 488 port select Remote from the Setup Menu then press Proceed The port selection screen will be disp
15. Microseal A is easily cut for use with fewer than 96 wells Microseal M rubber sealing mats are an economical means to seal 96 well microplates An array of 96 dimples on the mat helps orient it on the microplate and prevents the mat from sticking to the Hot Bonnet s heated lid The mats may be cleaned with sodium hypochlorite for reuse and are autoclavable Note After a hold at below ambient temperatures a ring of condensa tion may form in tubes above the liquid level but below the top of the sample block This is not a cause for concern since it occurs only at the final cool down step when thermal cycling is finished Adjusting the Hot Bonnet s Lid Pressure The pressure exerted by the inner lid of the Hot Bonnet must be manually adjusted to fit the sample vessels being used in a given reaction Once set the Hot Bonnet can be opened and closed repeatedly without readjustment as long as neither the tube or microplate type nor the sealing method changes Any change in vessel type or sealing method requires readjust ment of the Hot Bonnet s lid Follow these steps to adjust the pressure exerted by the Hot Bonnet s inner lid 1 Make sure the block s wells are clean Even tiny amounts of extraneous Operation W material can interfere with the proper seating of a microplate or tubes which would prevent the inner lid from exerting uniform pressure on the loaded microplate or tubes 2 Open the Hot Bonnet Turn th
16. Protocols in a password protected folder cannot be edited renamed or deleted nor can new protocols be placed in the folder without the pass word Users without knowledge of the password can still run copy and view a program Note A password cannot be assigned to the lt MAIN gt folder To assign a password to a folder select Secure from the Files Menu then press Proceed A list of all folders in the machine will be displayed Select the desired folder other than lt MAIN gt then press Proceed The password assignment screen will be displayed Secure lt FOLDER1 gt New password _ Valid passwords consist of numbers up to four digits long For passwords of three digits or less press Proceed after the password has been en tered Passwords four digits long will be automatically accepted as soon as typed and the Main Menu will be displayed 8 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual 8 4 Passwords can be changed at any time Follow the steps described above to select the desired folder then press Proceed A screen asking for the old password will be displayed Secure lt FOLDER1 gt Old password _ Enter the old password then press Proceed The password assignment screen be displayed Enter the new password then press Proceed The new password will be assigned to the folder and the Main Menu will be displayed again Deleting a Folder A folder must be empty
17. SS Co Gs S OS E CS C5 GOO E Co C S E lt lt 5 GSS OO c C as CO COCO x Q O ca rr GD gt eH SD 5 SS S55 ry i SSS SS lw KAI S S 22 a eN SS OE Ia AR 2 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual Alpha Units Single Block Models 60V Alpha Holds 60 x 0 5ml tubes 96V Alpha Holds 96 x 0 2ml tubes or one 96 well microplate 192 Alpha Holds one 192 well microplate 384 Alpha Holds one 384 well microplate or one 96 well microplate Dual Block Models 30 30 Dual Alpha Holds 2 x 30 x 0 5ml tubes 30 48 Dual Alpha Holds 1 x 30 x 0 5ml tubes and 1 x 48 x 0 2ml tubes 48 48 Dual Alpha Holds 2 x 48 x 0 2ml tubes or half plates Slide Block Twin Tower Holds 2 x 16 standard slides E H all DAS Power Bonnet Accessory Permits remote control of Alpha unit lid opening avail able for Alpha models 96V and 384 2 4 Specifications Thermal range Accuracy Thermal homogeneity Ramping speed Sample capacity Line voltage Frequency Power Fuses Displays Ports Memory Weight Size Layout and Specifications M 5 to 105 C but no more than 30 C below ambient temperature 0 3 C of programmed target 90 C NIST traceable 0 4 C well to well within 30 seconds of arrival
18. address to 0 Give the other machines in the network any number from 1 to 30 Press Cancel to delete a number and enter a new one Running a Protocol on Networked Machines To run a protocol on networked machines use the controller to select a protocol see Choosing a Stored Protocol to Run chapter 5 then press Proceed The network screen for the controller will be displayed Run 2 STEP on Block Local 96V The protocol will be identified on the top line 2 STEP in the example above The single asterisk in the upper right hand corner indicates that a single block Alpha unit is loaded into the controller Local indicates that this is the controller s network screen and 96V indicates the type of Alpha unit loaded into the controller If a dual block Alpha unit is loaded into the controller press Proceed until the letter designating the desired block is displayed after the aster isk in the upper right hand corner of the screen The letter A designates the right hand block and the letter B the left hand one e g A B To run the protocol on the controller s block press Proceed One or more screens for setting up the protocol s temperature control method will be displayed see Running a Protocol chapter 5 When these screens have been worked through the protocol will begin running To run the protocol on a different machine s block press Block until the screen identif
19. allows a progressive lengthening or shortening of hold times each time the step is executed in a GoTo cycle e Beep modifies a temperature step or a ramp step causing the ma chine to beep when a specified target temperature is reached Entering an Increment Option To enter an increment option select Inc from the Options Menu for a temperature step then press Proceed The first Inc screen will be dis 6 15 MW DNA Engine amp DNA Engine Tetrad Operations Manual 6 16 played Enter CUSTOM1 1 92 0 for 0 30 C cycle The temperature step being modified appears on the second line of this screen The plus sign on the third line means that the screen is set up to enter a progressive increase in temperature per cycle Press to switch to a minus sign allowing entry of a progressive decrease in temperature Press Cancel to change back to a plus sign Enter the numerical value of the temperature increase or decrease 1 2 is used in the example below Enter CUSTOM1 1 92 0 for 0 30 C cycle 1 2 Press Proceed The Inc value just entered will move to the third line of the screen and a confirmation menu will be displayed on the last line Enter CUSTOM1 1 92 0 for 0 30 1 2 per cycle OK _YES No Option Select one of the displayed choices then press Proceed e Yes accepts the Inc value and displays the Enter Menu again Use the Enter Menu to enter the next ste
20. before it can be deleted After all programs have been moved or deleted from the folder select Delete from the Files Menu then press Proceed A list of all folders in the machine will be displayed Select the folder to be deleted then press Proceed A confirmation screen will be displayed Delete lt FOLDER1 gt Delete empty folder YES No Select Yes then press Proceed The folder will be deleted and the Main Menu will be displayed To cancel the deletion press Cancel or select No and press Proceed Copying a Program The copy utility copies a program and gives the copy a new name Cop ies can be placed in the original folder or a new one To copy a program select Copy from the Files Menu then press Pro ceed Locate the program to be copied see Locating a Stored Program p 8 2 then press Proceed If more than one folder of programs is present in the machine a screen allowing you to specify the folder the program will be copied to will be displayed Copy CUSTOM1 to _ lt FOLDER1 gt lt FOLDER2 gt lt FOLDER2 gt Using the Utilities Mi Select a folder to copy the program to then press Proceed A naming screen will be displayed Copy CUSTOM1 to Name the copied program see Naming the Program chapter 6 then press Proceed The program will be copied to the specified folder un der the new name and the Main Menu will be displayed Renaming a Pr
21. dimer production by designing primers with no 3 self complementarity Reoptimize magnesium concen tration and annealing temper ature to maximize desired pro duct and minimize primer dimers Program a hot start into the protocol Check concentratons of compo nents May need to reoptimize magnesium concentration Reoptimize annealing temperature Use List utility to check proto col s temperature control method temperatures and times Check extraction and purifi cation protocols Add additional purification steps if necessary Check nucleic acid preparations by gel electrophoresis Problem No reaction products obtained Cause Wrong protocol used Protocol contains a wrong value Reaction component omitted from mixture Denaturation temperature too low Annealing temperature too high for primers Wrong temperature control method used Probe failed causing machine to run protocol under calculated control Probe not filled with correct amount of oil Reaction mix contains an inhibitor e g heme from blood Reaction vessels not making good thermal contact with sample block Troubleshooting Mi Action Re run reaction using correct protocol Use List utility to check proto col s temperature control meth od temperatures and times Check reaction assembly proto col ensuring that mixture con tains appropriate components in correct concentrations Use g
22. diminishes thermal coupling of sample vessels to the block Caution Do not pour any cleaning solution into the block s wells and then heat the block in an attempt to clean it Severe damage to the block the heated lid and the chassis will result Cleaning the Air Vents Clean the air intake and exhaust vents with a soft bristle brush a damp cloth or a vacuum cleaner whenever dust is visible in them The air in take vents are located on the bottom lower front edge and back of the machine the air exhaust vents are located on both sides see figs 2 1 2 3 and 2 4 If these vents become clogged with dust and debris airflow to the Alpha unit s heat sink is hampered causing performance problems re lated to overheating The air intake vents are particularly likely to collect dust since their holes are much smaller than those of the air exhaust vents Y Tip To prevent problems with overheating institute a regular pro gram of checking for dust buildup particularly for robotics installations Maintenance Cleaning Radioactive or Biohazardous Materials Out of the Block When cleaning machines that have been running radioactive or biohaz ardous reactions consult your institution s radiation safety officer or biosafety officer regarding cleaning methods monitoring and dispos ing of contaminated materials Changing the Fuses The circuits in the DNA Engine are protected by two fuses 6 3A fast acting 5 x 20mm Whe
23. fig 4 1A 5 Push the handle down until it is completely vertical fig 4 1B firm pressure may be required A definite click signals that the Alpha unit s connectors have mated with the DNA Engine s connectors When the handle is in the down position the Alpha unit is locked into place Installing an Alpha unit 4 4 Operation W Removing an Alpha Unit 1 Turn the DNA Engine off see the Caution below 2 Pull upward on the handle When the lock releases you will hear a click and the Alpha unit will slide a little toward the back of the DNA Engine The electrical connectors of the Alpha unit and the DNA En gine are now disengaged so there is no danger of electrical shock 3 Slide the Alpha unit toward the rear of the DNA Engine about 3cm 4 Grasp the front and back edges of the Alpha unit and lift it out of the machine Caution Do not insert or remove an Alpha unit with the DNA Engine turned on electrical arcing can result Read the safety warning in appendix A regarding electrical safety when inserting or removing an Alpha unit Opening an Alpha Unit Grip the front edge of the top lever of the Hot Bonnet as shown in figure 4 2A and pull upward firmly The top lever will pop open to reveal the entire thumbwheel fig 4 2B Continue pulling upward to open the Hot Bonnet The Hot Bonnet will tip backward revealing the entire block Caution Do not pull on the thumbwheel to open the unit This can d
24. screws Operating the DNA Engine Tetrad Turning the DNA Engine Tetrad On First make sure Alpha units are installed in all four quadrants An in terlock switch prevents the DNA Engine Tetrad from operating if fewer than four Alpha units are installed Turn the DNA Engine Tetrad on with the Power key located at the lower right of the control panel Operating Alpha Units Alpha unit installation removal and general operation are the same as for the DNA Engine see chapter 4 Running Protocols on the DNA Engine Tetrad Running Separate Protocols by Block Choose a protocol see chapter 5 A screen identifying the protocol 2 STEP in the examples below the Alpha unit type 96V and the quad rant number of the block the protocol will run on 1 will be displayed Run 2 STEP on 1 Block 96V If a dual block Alpha unit is present in the identified quadrant an A or a B will appear after the quadrant number e g 1A Protocols are initially assigned to the first block available in the DNA Engine Tetrad in numerical order of quadrants To run the protocol on a different block press Block until the quadrant number of the desired block is displayed in the screen s upper right hand corner The status light of the selected block will flash When a single block Alpha unit is selected only the A light flashes When a dual block Alpha unit is se lected either the A or the B light flashes depending on the block c
25. scroll to the step you want to edit Reviewing a gradient program After the protocol has been run the Program Complete screen will be dis played Press the Proceed key and the next screen will display a re view of the gradient well temperatures Gradient Review Column 1 50 0 left Column 2 50 59 Column 3 51 5 Column 4 53 2 Column 5 55 52 Column 6 58 4 Column 7 61 8 Column 8 64 6 Column 9 66 82 Column 10 68 4 Column 11 69 6 Column 12 70 0 right Scroll up and down the screen using the Select keys When scrolling the Gradient Review line will be held constant while the lower 3 lines can be manipulated up or down The temperatures displayed will be for each of the twelve wells along the long axis of the sample block These temperatures will be predicted from an algorithm as a steady state tem perature of the block and the sample You may then return to the main menu by pressing the Proceed key Using the gradient calculator The gradient calculator predicts for a given gradient the temperatures for each of the twelve wells along the long axis of the sample block These temperatures will be predicted from an algorithm as a steady state tem perature of the block and the sample To use the gradient calculator select the List command from the main menu and then press the Pro ceed key You will see a List screen Creating Programs W List Program grams _GRADIENT CALCULATOR
26. siii ii B 1 Appendix C How a Switching Power Supply Works ooooccionnncnncncnnonccnocononannnnnnnanoss C 1 Appendix D Shipping Instructions for US Residents 00 0 0 eeeseseeeseeceeeeteeeeeeeeeeees D 1 APPENA E Wat 3 saxicvngon us eau E T EET T E 1 Appendix F Factory Installed Protocol F 1 Documentation Conventions Typographic Conventions The names of keyboard keys are set in sans serif type and encased in double angle brackets Example Proceed Items in programming menus are italicized Example Select Edit from the Main Menu Graphic Conventions Terminology The programming screens displayed in the LCD window are represented by a box containing four lines of text Example Run 2 STEP 1 92 0 for 0 05 Cycle 1 Calc 65 0 A programming option is termed selected when the cursor is positioned in front of it Use the Select keys see fig 2 2 to move the cursor In some screens selected items are also displayed in all capital letters The DNA Engine Introduction Meet the DNA Engine 1 2 Using This Manual 1 2 Important Safety Information 1 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual Meet the DNA Engine Thank you for purchasing an MJ Research PTC 200 DNA Engine De signed by a team of molecular biologists and engineers the DNA Engine will meet your needs for a versatile easy to use reliable and compact programmable thermal cycler Interchangeable samp
27. step involves an option as well as a program step write both names down on the same line Finally write the End step at the bottom of the list pro grams will not run without this step Number the lines 1 through N where N is the final End line Using the GoTo Step to Write Short Programs The GoTo step allows programs of many repetitious steps to be short ened to just a few lines When the program encounters a GoTo step it returns to a specified step repeats that step and repeats all steps that follow back to the GoTo step When the program has returned or cycled back to the step a specified number of times the program moves on to the step that follows the GoTo step For example consider a basic cycle sequencing protocol consisting of 30 repeats of a denaturation and an annealing extension step Rather than listing all 60 steps use a GoTo step to design a short easy to enter pro gram Raw program DNA Engine program 1 92 for 30 sec 1 92 for 30 sec 2 60 for 3 min 2 60 for 3 min 3 92 for 30 sec 3 GoTo step 1 29 times i e 4 60 for 3 min cycle back to step 1 and repeat 5 92 for 30 sec steps land 2 29 times 6 60 for 3 min 5 End 7 92 for 30 sec continues for total of 60 lines Choosing a Temperature Control Method The DNA Engine can control block temperature in three possible ways each of which has different implications for the speed and accuracy of sample heating e Calc
28. the DNA Engine Chapter 9 describes how to network and remotely operate the DNA Engine and DNA Engine Tetrad Introduction W e Chapter 10 explains the proper maintenance of the DNA Engine e Chapter 11 offers troubleshooting information for the DNA Engine e Chapter 12 describes the operation of the DNA Engine Tetrad e Chapter 13 describes the operation of the RAD 0200 Remote Alpha Dock accessory Important Safety Information Safe operation of the DNA Engine or DNA Engine Tetrad begins with a complete understanding of how the machine works Please read this en tire manual before attempting to operate the DNA Engine or DNA En gine Tetrad Do not allow anyone who has not read this manual to oper ate the machine The DNA Engine and DNA Engine Tetrad can generate enough heat to inflict serious burns and can deliver strong electrical shocks if not used according to the instructions in this manual Please read the safety warn ings and guidelines in appendix A and exercise all precautions outlined in them 1 3 Front View 2 2 Control Panel 2 2 Back View 2 3 Bottom View 2 3 Alpha Units 2 4 Single Block Models 2 4 Dual Block Models 2 4 Slide Block 2 4 Power Bonnet Accessory 2 4 Specifications 2 5 Gradient Specifications 2 6 Layout and Specifications 2 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual Front View Figure 2 1 Thumbwheel Alpha unit lid closed FS eS fs GU
29. the outside after thermal cycling Please be certain that you are using the 4 15 Operation W appropriate detection methods and cleaning procedures for this isotope Consult your radia tion safety officer for his or her recommendations If mild cleaning agents do not remove radioactivity harsher cleaners may be used Users have suggested the detergent PCC 54 Pierce Chemical Co Rockford Illinois Pierce Eurochemie B V Holland Micro Cleaning Solution Cole Parmer Niles Illinois and Dow Bathroom Cleaner avail able in supermarkets Caution Harsh cleaning agents are corrosive to aluminum and must never be used on bare aluminum blocks MJ ResearcH blocks are anodized so they have a protective coating of aluminum oxide Still harsh agents such as those above must be thoroughly rinsed away within a few minutes of application or the anodization will degrade 4 16 Appendix 4 C Using Silicone Oil in the Probe Tube The following light and heavy silicone oils may be used instead of mineral oil as the representa tive sample in a probe tube e Dow Corning 200 light silicone oil dimethypolysiloxane Sigma DMPS 5X Density 0 97g ml Viscosity 50cs Volume to use 1 7 x volume of buffer in individual sample tube plus one volume of oil overlay Dow Corning 200 heavy silicone oil dimethypolysiloxane Sigma DMPS V Density 0 97g ml Viscosity 5cs Volume to use 2 7 x volume of buffer in individual sample tube p
30. to Protocols 11 6 Problems Related to Environmental Conditions Setup and Maintenance 11 8 11 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual Error Messages Error Message A C Power Failed Cycle XXX Step X Recovered at XX X Block Overheated Return for Service Block Sensor Fault Left Ignored Center Ignored Right Ignored Block Sensor Fault Program Terminated 11 2 Note The DNA Engine Tetrad software is highly sensitive with re spect to block and heat sink errors When such errors messages occur try restarting the protocol If the message fails to reappear proceed as usual Cause and Result Displayed when a machine run ning a protocol has been turned off either intentionally or due to a power outage and then turned on again On machines with software ver sions below 1 1M can happen when running calculated control protocols with target temperatures of 103 105 C As the calculated control algorithm tries to hold samples at such high tempera tures it can heat the block to tem peratures in excess of the Alpha unit s maximum allowable tem perature 107 5 C triggering an automatic Alpha unit shutdown Sensor malfunction allows base to heat block over its maximum al lowable temperature triggering an automatic Alpha unit shut down Left center or right temperature sensor in Alpha unit is not work ing properly so machine is now ignoring it and relying on remain
31. 4 environmental requirements 13 3 installation 13 5 operation 13 6 packing checklist 13 3 power supply requirements 13 4 Robotics installation requirements 3 5 Index M S Safety general instructions 1 3 guideline for safe use A 1 warnings A 1 Sample vessel selection chart 4 14 Sample vessels ensuring good thermal contact 4 10 loading into block 4 9 ensuring good thermal contact 4 9 sealing Reason for 4 7 with Hot Bonnet and Caps Film 4 8 with oil or wax 4 7 selection 4 6 0 2 ml tubes 4 6 0 5 ml tubes 4 6 microplates 4 6 Use of oil to improve thermal contact with block 4 10 Self test 4 2 Setting machine up See also Alpha units installing Shipping instructions E 1 E 1 Specifications 2 5 DNA Engine Tetrad 2 5 Switching power supply D 1 T Temperature control methods block control 6 3 6 4 calculated control 6 3 6 4 choice of calculated control over probe control 4 11 probe control 6 4 Thermoelectric unit C 1 U Utilities file assigning password to folder 8 3 8 4 copying program 8 4 creating folder 8 3 deleting a program 8 5 Deleting folder 8 4 moving a program 8 5 renaming programs 8 5 List 8 6 In 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual viewing a program in LCD window 5 6 list printing a program 8 7 setup 8 7 choosing a printer port 8 8 choosing a remote port 8 7 choosing a temperature control method for the Hot 8 9 determining software version number 8 9 In 4 DECLARATION O
32. A Engine Tetrad harnesses four DNA Engines in a single com pact machine Four Alpha units of any type can be loaded into the DNA Engine Tetrad base one unit into each quadrant of the base The same factory installed and custom protocols can be run on the DNA Engine Tetrad as on the DNA Engine Programs can be run independently on any one Alpha unit or simultaneously on all of them The DNA Engine Tetrad s control panel Figure 12 1 Layout of quadrants is nearly identical to that of the DNA En gine Additional features include a Back of Tetrad Power key and an array of lights indi cating the selection status of the blocks in the base s quadrants There is an Aand a B light for each quadrant since each Al pha unit can contain up to two blocks The arrangement of the status indicator lights matches the arrangement of the quadrants in the base when the DNA Engine Tetrad Front of Tetrad is viewed from above fig 12 1 Quadrant 1 Quadrant 2 y JUDIPONO JUDIPONE The DNA Engine Tetrad s power supply has been placed in a separate housing that is attached by cables to the base which keeps the base com pact and makes it easier to use in robotics installations The power sup ply uses MOSFET transistors to efficiently convert AC input power into the high current low voltage DC power that the Tetrad requires see ap pendix D for a short explanation of this technology Up to 15 DNA Engine Tetrads and DNA Engines can be net
33. A regarding electrical safety when inserting or removing an Alpha unit Environmental Requirements Ensure that the area where the DNA Engine is installed meets the follow ing conditions for reasons of safety and performance e Nonexplosive environment e Normal air pressure altitude below 4000m 3 2 Installation M e Ambient temperature 5 31 C e Relative humidity up to 90 e Unobstructed access to air that is 31 C or cooler see below e Protection from excessive heat and accidental spills Do not place the DNA Engine near such heat sources as radiators and protect it from danger of having water or other fluids splashed on it which can cause shorting in its electrical circuits Power Supply Requirements The DNA Engine requires 100 240VAC 50 60Hz and a grounded out let The DNA Engine can use current in the specified range without adjustment so there is no voltage setting switch Power cords for outlets other than the US 120V outlet may be purchased from computer stores since they are also used for most desktop computers and printers and meet international standard IEC 320 The power cord must be rated to carry at least 10A at 125V or 250V depending on the voltage available in your nation The quality of the power cord can be further en sured by making certain it is inscribed with the trademark of UL CSA TUV VDE or another national testing agency Note Do not cut the supplied 120V power cord and attach a di
34. Change All the Values in the Step Being Entered Repeatedly press Cancel Each time you press the key the cursor will move backward through the values for the step deleting the values as it moves When all values for the step have been deleted enter new val ues or press Cancel one more time to display the Enter Menu for the step At this point you can change the step to a different type To Change Values for Earlier Steps in the Program Repeatedly press Cancel until the Enter Menu for the step you are working on is displayed Press Cancel one more time to display a list of all steps entered so far The list will show the step number and the values for each step including any options chosen The last entered steps will be displayed first Enter CUSTOM1 3 55 0 for 0 30 72 0 for 1 00 Creating Programs Mi At this point you can choose to change selected values in program steps to add an option to enter Edit mode or to delete the entire program Press the Select keys to scroll through the program The left Select key moves the cursor backward through the steps and the right Se lect key moves it forward e To change selected values or add an option To change a value move the cursor to it then press Cancel Enter the new value then press Proceed If you change your mind press Cancel again the de leted value will be restored To add an option to a step move the cursor to the st
35. Deleting a Step To delete a step select Delete from the Edit Menu then press Proceed The step will immediately be deleted and the program being edited will be displayed again minus the deleted step To cancel a deletion see Cancelling Editing Changes p 7 5 Note Be careful when using Delete Once a step has been deleted it cannot be recovered without abandoning all editing changes that have been made in the program This could be inconvenient if the program has been extensively edited Adding an Option To add an option to a step select Option from the Edit Menu then press Proceed The option menu will be displayed for the step Add the desired option to the step see Modifying a Program Step with the Options chapter 6 When the option is complete select Yes from the confirmation menu then press Proceed The steps of the pro gram being edited will be displayed again with the new option ap pearing in the list Saving an Edited Program 7 4 To save an edited program use the right Select key to scroll to the End step of the program Position the cursor on the number for the step then Editing Programs W press Proceed A line allowing the editing session to be ended will be displayed on the last line of the screen Edit CUSTOM1 2 End Step _END Insert e End saves the changes and displays the Main Menu This ends the editing session e Insert allows another step to be add
36. F CONFORMITY TO APPLY CE MARK MJ RESEARCH INC hereby declares that the MJ RESEARCH PTC 200 DNA Engine thermal cycler conforms to the following directives and standards APPLICATION OF COUNCIL DIRECTIVES 89 336 EEC amp 73 23 EEC STANDARDS TO WHICH CONFORMITY IS DECLARED EN55011 EN50082 1 EN61010 1 EMC for industrial scientific medical equipment EMC for generic immunity compliance to low voltage directive respectively TYPE OF EQUIPMENT Scientific Equipment for Laboratories MODEL NUMBER PIC 200 YEARS OF MANUFACTURE 1997 and onward MJ RESEARCH INC as manufacturer of the equipment described above certifies that this model has been tested and conforms to the applicable Directives and Standards of the European Union EU as described above Test data to verify this conformity are available for inspection at either MJ RESEARCH INC or Finnzymes Oy at the addresses listed below The instrument in question carries the CE mark on the nameplate MANUFACTURER S OFFICIAL A od Robert A lovanni Senior Electrical Engineer MJ RESEARCH INC 590 Lincoln Street Waltham Massachusetts 02451 UNITED STATES OF AMERICA Tel 617 923 8000 Fax 617 923 8080 Date 8 5 99 DECLARATION OF CONFORMITY MJ RESEARCH INC hereby declares that the MJ RESEARCH PTC 225 DNA Engine Tetrad thermal cycling system conforms to the following directives and standards APPLICATION OF COUNCIL DIRECTIVES 89 336 EEC
37. Japan The DNA Engine Tet rad can use current in the specified range without adjustment so there is no voltage setting switch Environmental Requirements The requirements for safe operation of the DNA Engine Tetrad are the same as those of the DNA Engine see chapter 3 Air Supply Requirements The air supply requirements for the DNA Engine Tetrad are the same as those of the DNA Engine see chapter 3 Robotics Installation Requirements Robotics installations of DNA Engine Tetrads require special attention to airflow and air temperature Typically in these installations DNA En gine Tetrads and other thermal cyclers are crowded into a small area along with other heat generating equipment Overheating can quickly occur when many of these machines are operating at once unless pre ventive measures are taken Follow the procedures described in chapter 3 to ensure adequate airflow and an air intake temperature of 31 C or cooler Air intake temperature must be verified by measurement If none of the troubleshooting proce dures outlined in chapter 3 are sufficient to ensure cool enough air install ductwork to the front and rear air exhaust vents to remove exhausted air Attach ductwork using the screws to the right and left Information Specific to the DNA Engine Tetrad Mi of the air exhaust vents on the front and back of the DNA Engine Tetrad fig 12 2 These screws may be replaced with longer ones as long as they are M4 x 0 7
38. NA Engine amp DNA Engine Tetrad Operations Manual 6 6 Initiating the Program To initiate a new program select Enter from the Main Menu then press Proceed A naming screen will be displayed Naming the Program Name the program an eight character word consisting of any combina tion of letters Roman and Greek numbers punctuation marks or Japa nese Katakana Press the right Select key to scroll forward and the left Select key to scroll backward through the alphabets and characters available which are presented in this order Roman alphabet selected Greek letters punctua tion marks numbers To access the Japanese Katakana syllabary press the key A second press of returns the machine to Western characters When the character needed is displayed next to Name press Proceed The character will be accepted and the cursor will move one space to the right Numbers and dashes may also be inserted by pressing the corre sponding keys on the keypad When the name is complete press Proceed once to accept the last char acter and again to accept the whole name If the name is already in use for a program a screen saying Name In Use will be displayed If this happens press Proceed then enter a different name Choosing a Temperature Control Method When the name has been entered CUSTOM1 is used in the following and all succeeding examples a screen requesting selection of a te
39. Never open the DNA Engine Tetrad base you could receive a serious electrical shock Opening the base will also void your warranties Figure 12 6 Quadrant assignments for fuses 12 10 Quadrant 4 Quadrant 1 Quadrant 3 Quadrant 2 Part Ill Accessories 1 3 The Remote Alpha System About the Remote Alpha Dock System 13 2 Packing Checklist 13 3 Requirements 13 3 Installation 13 5 Operation 13 6 Dock 13 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual About the Remote Alpha Dock System Figure 13 1 13 2 The Remote Alpha Dock system is designed to add flexibility to the in stallation and operation of the MJ Research PTC 200 DNA Engine and PTC 225 DNA Engine Tetrad The system allows Alpha units to be placed at a distance from the PTC 200 or PTC 225 base enabling more efficient use of space and facilitating robotic operation The basic system the RAD 0200 comprises a Dock Connector which mounts in the base and a Re mote Alpha Dock into which the Alpha units are mounted The RAD 0201 contains in addition a fan power supply and three daisy chain series cables allowing the cooling fans for up to four Remote Docks to run off a single AC outlet Additionally the fan power supply along with the cables to run up to four Remote Alpha Dock fans from a single ther mal cycler base can be purchased separately as RPS 0200 The Remote Alpha Dock with Alpha unit mounted The Remote Al
40. No then press Proceed The protocol will begin running Running Protocols W Setting Up a Probe Control Protocol A single screen asking about use of the heated lid will be displayed Se lect Yes or No then press Proceed If the probe is installed the probe verification screen will be displayed Run PROBE1 PROBE control Verify probe ready Ensure that the probe is correctly installed see Loading and Connecting the Probe chapter 4 then press Proceed The protocol will begin running If the probe is not installed a warning message will be displayed RUN PROBE1 CALC control Probe not present This screen says that since the probe is missing the DNA Engine will run the protocol using calculated control if Proceed is pressed now To avoid this press Cancel install the probe and set the protocol up again When the probe verification screen is displayed press Proceed to be gin the protocol run Reading the Runtime Screen During a protocol run a runtime screen will be displayed Run 2 STEP 1 92 0 for 0 05 Cycle 1 Calc 68 0 The screen lists the protocol name 2 STEP in the example above proto col step that is running 1 cycle number 1 method of temperature control Calc and the block temperature for block control protocols the calculated sample temperature for calculated control protocols 68 0 and the probe temperature for probe control protocols When
41. Record of Programs 6 20 Occasionally in the course of repairing a defective DNA Engine it is necessary to replace the chip that stores all custom user protocols To avoid losing your protocols in such an event always maintain an up to date record of them Protocols may be printed out using the List utility see chapter 8 stored on a computer using the DNA Engine Driver Soft ware or written down in a notebook Editing Programs Editing a Stored Program 7 2 Initiating Editing 7 2 Editing the Program 7 3 Inserting a New Step 7 4 Deleting a Step 7 4 Adding an Option 7 4 Saving an Edited Program 7 4 Cancelling Editing Changes 7 5 7 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual Editing a Stored Program The DNA Engine s editing tools available through Edit on the Main Menu make it easy to extensively edit stored programs by Changing individual values in program steps Adding new steps Deleting steps and Adding options to temperature and ramp steps Note The editing tools do not include a renaming function To re name a program see Renaming a Program chapter 8 Initiating Editing To initiate editing select Edit from the Main Menu then press Pro ceed One of two types of screen will be displayed depending on whether your programs have been stored in the lt MAIN gt folder or in custom folders If all programs have been stored in lt MAIN gt The screen will list the
42. STANDARDS TO WHICH CONFORMITY IS DECLARED EN55011 EN50082 1 EMC for industrial scientific medical equipment amp EMC for generic immunity TYPE OF EQUIPMENT Scientific Equipment for Laboratories MoDEL NUMBER PTC 225 System which includes one PTC 225 base unit four PTC 200 225 Alpha sample block heat pump subassemblies and one AT amp T RM2000 Power Supply YEAR OF MANUFACTURE 1996 MJ RESEARCH INC as manufacturer of the system described above certifies that this model has been tested and conforms to the applicable Directives and Standards of the European Union EU as described above Test data to verify this conformity are avail able for inspection at either MJ RESEARCH INC or Finnzymes Oy at the addresses listed below The instrument in question carries the CE96 mark on the nameplate MANUFACTURER S OFFICIAL y ae Robert A lovanni Senior Electrical Engineer MJ RESEARCH INC 590 Lincoln Street Waltham Massachusetts 02451 UNITED STATES OF AMERICA Tel 617 923 8000 Fax 617 923 8080 Date 8 5 99 EU EU ee ee O O A Fy F NA F Na cy a pa pa pa po F a O A OQ A wa E OQ f g Loy Loy Loy F a A g NA F NA a cy a a A OQ eq OQ T a OQ E O os Ey to gt y to a q tdo a 4 a A 4 MJ RESEARCH INC on e Y Rp ONAN a Manufactory a a T AQA A A 590 Lincoln Street Yoo OO y o l Waltham Massachusetts 02451 USA o pan P o E Product Information 888 735 8437 7 E a i Cust
43. Setup The DNA Engine is now ready to execute programs Understanding the Main Menu 4 2 The Main Menu is the common access point to all programming and machine configuration screens e Run Executes a program e Enter Allows new programs to be entered e List Accesses utilities that display or print a program s steps e Edit Allows modification of stored programs e Files Accesses file management utilities Setup Accesses machine and networking configuration screens Operation W Using the Control Panel The control panel see fig 2 2 includes operation keys status indicator lights an LCD window for displaying programming and machine status text and a numeric keypad for entering values into programs Operation Keys e Select keys left and right arrows Move the cursor one space or op tion to the left or right in the LCD window display time and cycle information during a protocol run e Proceed Accepts a selected menu or screen option during a protocol run advances the program to its next step e Cancel Terminates a running protocol during program creation or editing cancels the last entry Stop Terminates a running protocol e Pause Pauses a protocol during execution accesses Japanese Katakana syllabary Instant Incubate Initiates a program that sets up the DNA Engine as a simple incubator e Block Selects a different block when using a dual block Alpha unit switches betwee
44. T Sono back moy be ATENG labs pewanab mien LWA Lp Yi A O bod Sons O g lt Mean Cooling P Control panel bd Lily Air exhaust vents also on other side PIC Peltier Th er eA 7 Oder Air intake vents Control Panel Figure 2 2 Block key CAUTION Sample blocks may be very hot ATTENTION Les blocs peuvent tre tres chauds AO BlockStaws O B LCD window e Heating e Cooling Block status lights Instant incubation key 4 Instant Incubate Left and right selection keys Power light A Proceed key Proceed MJ RESEARCH Cancel PTC 200 Peltier Thermal Cycler Y Cancel key 2 2 Layout and Specifications M Back View Figure 2 3 Air exhaust vents Alpha unit also on other side SJ i nl I Alpha unit handle Air intake vents Power cord jack IEEE 488 port RS 232 port Parallel printer port Bottom View Figure 2 4 Air intake vents Front f AL orooro al S S S JCS SS cSSc gt Sa C3535 GS Sac Co G5 Cn CSD G 4 CS 2 SEO 44 G5 G5 E 4 C5 0 OOO Co HS Gs GOO Co Cn GS GD GS a Co CH GO U CO D Co 4 8 GSO 1 D J Coa CS SSS CSCS Cy CSS AAA Co UU R S OO Y CS Y S SO O lt lt Cn AY S O O lt lt Cn Gs 55 GS O Co CH GS GOES Co Cn WS GS GSI a SS ee es ee CS GS 3 CSC 444 CH G5 GS GS Co OE Co S GS D Ce CS G5 UE
45. _CUSTOM1 CUSTOM2 QUIKSTEP 2 STEP 3 STEP EXTEND Select the desired program from this list e If programs have been stored in custom folders The screen will list all the folders residing in the machine Copy lt MAIN gt lt FOLDER1 gt lt FOLDER2 gt lt FOLDER3 gt Select the folder containing the program then press Proceed A list of all programs in the folder will be displayed Select the desired pro gram then press Proceed File Utilities Use these utilities available from the Files Menu to accomplish the fol lowing tasks e Create folders to store programs in e Assign passwords to folders e Copy delete rename or move programs To display the Files Menu select Files from the Main Menu then press Proceed Files _COPY Delete Move Rename Folder Secure 8 2 Using the Utilities Mi Creating a Folder The DNA Engine s memory can hold up to 11 folders including the lt MAIN gt folder New programs are placed in the lt MAIN gt folder by de fault unless a different folder is specified To create a folder select Folder from the Files Menu then press Pro ceed A naming screen will be displayed New Folder Name the folder using the instructions found under Naming the Pro gram in chapter 6 then press Proceed The name will be assigned to the new folder the folder will be stored and the Main Menu will be dis played Assigning a Password to a Folder
46. air intake vents following procedure on p 3 3 If higher than 31 C use table 3 1 to troubleshoot and remove cause s Remove light collections with damp cloth Vacuum out heavy collections The DNA Engine Tetrad Information Specific to the DNA Engine Tetrad Meet the DNA Engine Tetrad 12 2 Specifications 12 2 Layout 12 3 Front View 12 3 Control Panel 12 3 Back View 12 4 Bottom View 12 4 Unpacking and Installing the DNA Engine Tetrad 12 5 Packing Checklist 12 5 Installation 12 5 Connecting the Cables 12 5 Power Supply Requirements 12 6 Environmental Requirements 12 6 Air Supply Requirements 12 6 Robotics Installation Requirements 12 6 Operating the DNA Engine Tetrad 12 7 Turning the DNA Engine Tetrad On 12 7 Operating Alpha Units 12 7 Running Protocols on the DNA Engine Tetrad 12 7 Running Separate Protocols by Block 12 7 Running the Same Protocol on All Blocks 12 8 Running an Instant Incubation 12 8 Cancelling Protocols 12 8 Programming the DNA Engine Tetrad 12 9 Networking the DNA Engine Tetrad 12 9 Software Required for Networking 12 9 Operating Networked DNA Engine Tetrads 12 9 Using the Utilities for the DNA Engine Tetrad 12 9 Troubleshooting the DNA Engine Tetrad 12 9 Maintaining the DNA Engine Tetrad 12 10 Cleaning 12 10 Changing the Fuses 12 10 12 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual Meet the DNA Engine Tetrad Specifications 12 2 The DN
47. amage the Hot Bonnet s closing mechanism Figure 4 2 Opening an Alpha unit 4 5 MW DNA Engine amp DNA Engine Tetrad Operations Manual Closing an Alpha Unit Press down on the top lever The lever will close down over the thumbwheel as the Hot Bonnet closes down over the sample block A click signifies that the Hot Bonnet s latch has engaged Selecting the Correct Sample Vessel 4 6 The DNA Engine s wide variety of interchangeable Alpha units affords you great scope in choosing sample vessels Keep in mind that differ ences in tube and plate composition and wall thickness among the many brands available can affect reaction performance Protocols may require some adjustment to ensure optimum performance when using a new vessel type MJ RESEARCH offers a full range of tubes and microplates manufactured to the specifications of each type of Alpha unit to ensure a precise fit See chapter appendix 4 A for a complete list 0 5 ml Tubes Make sure thick walled 0 5 ml tubes fit the wells snugly Since these tubes were originally designed for centrifuges some brands may not fit tightly in thermal cycler wells Thin walled 0 5 ml tubes were specifically de signed for thermal cycling and the higher quality brands provide a good and consistent fit MJ RESEARCH provides thin and thick walled 0 5 ml tubes designed for precise block fit 0 2 ml Tubes All types of thin walled 0 2 ml tubes may be used MJ Research sells high quali
48. are crowded 3 4 Possible Remedies Provide more space around machine or adjust room ventilation Adjust air conditioning to lower ambient air temperature Move machine away from or protect machine from such heat sources as radiators heaters other equipment or bright sunlight Arrange machines so that warm exhaust air does not enter intake vents Installation M Requirements for Robotics Installations Robotics installations require special attention to airflow and air tempera ture Typically in these installations DNA Engines and other thermal cy clers are crowded into a small area along with other heat generating equip ment Overheating can quickly occur when many of these machines are operating at once unless preventive measures are taken Follow the procedures described above to ensure adequate airflow and an air intake temperature of 31 C or cooler Air intake temperature must be verified by measurement Do not use oil to thermally couple sample vessels to the blocks of ma chines in a robotics installation Oil makes plates difficult to remove 3 5 ET operation Turning the DNA Engine On 4 2 Understanding the Main Menu 4 2 Using the Control Panel 4 3 Operation Keys 4 3 Status Indicator Lights 4 3 Using the Data Ports 4 3 Operating Alpha Units 4 4 Installing an Alpha Unit 4 4 Removing an Alpha Unit 4 5 Opening an Alpha Unit 4 5 Closing an Alpha Unit 4 6 Selecting the Correct Sample Vessel 4
49. area e Jefe peje EE EE e CE pupnezzvamegins MPa e fe fe po po conor enemigas ONS Papers msm e fete jojo porem mn e ef fofe pamase CT fe fe panties ways IET pate wap in wal Ta _ a 0 5 ml tubes w caps thick wall T6 1000 A IE Microseal M and P sealers are sized for 96 well blocks but can be used with fewer than 96 wells if the vessels are placed symmetrically in the block Microseal P pads are intended for use with Power Bonnet motorized lids Note All tubes and multiplate microplates are made from polypropylene plastic the optimal material for this application Concord microplates are made from polycarbonate plastic which is more prone to poor sealing and vapor leakage 4 14 Appendix 4 B Safety Warning Regarding Use Of S Nucleotides Some researchers have experienced a problem with radioactive contamination when using S in thermal cyclers This problem has occurred with all types of reaction vessels The Problem When S nucleotides are thermally cycled a volatile chemical breakdown product forms prob ably SO This product can escape the vessel and contaminate the sample block of a thermal cycler and possibly the air in the laboratory Contamination has been reported with microassay plates 0 2 ml tubes and 0 5 ml tubes 96 Well and 192 Well Polycarbonate Microplates These microplates present the largest risk of contamination Polycarbonate is so
50. at 90 C for most Alpha units see specifications for in dividual Alpha units Up to 3 C sec for all single and dual block Alpha units 1 2 C sec for the Twin Tower Varies with installed Alpha unit 100 240VAC rms no adjustment needed among volt ages within these ranges 50 60Hz single phase 850W maximum Two 6 3A 250V 5 x 20mm One 20 x 4 LCD alphanumeric display One 25 pin 8 bit parallel interface printer port One 9 pin RS 232 serial port for printer or remote use One IEEE 488 bidirectional general purpose interface bus 200 typical programs in up to 12 individual folders 7 6 9 3kg varies with installed Alpha unit DNA Engine Tetrad 29kg including Alpha units and power supply 24 x 35 x 17cm DNA Engine Tetrad 37 x 55 x 25cm 2 5 Gradient Specifications 96V Alpha module only Accuracy 0 4 C of programmed target at end columns 30 seconds after the timer starts for the gradient step NIST traceable Column uniformity 0 4 C well to well within column within 30 sec onds of reaching target temperature Calculator accuracy 0 4 C of actual well temperature Lowest programmable 30 C temperature Highest programmable 105 C temperature Temperature differential 1 24 C range Installation Packing Checklist 3 2 Setting Up the DNA Engine 3 2 Environmental Requirements 3 2 Power Supply Requirements 3 3 Air Supply Requirements 3 3 Ensuring an Adequate Air Supply 3 3 Ensurin
51. bris should be vacuumed out Turn the ma chine off prior to cleaning or vacuuming air vents Ensuring That Air Is Cool Enough Do not position two or more DNA Engines or other thermal cyclers so that the hot exhaust air of one blows directly into the air intake vents of another Make sure the DNA Engine receives air that is 31 C or cooler by mea suring the temperature of air entering the machine through its air intake vents Place the DNA Engine where you plan to use it and turn it on Try to reproduce what will be typical operating conditions for the machine in that location particularly any heat producing factors e g nearby equipment running window blinds open lights on Run a typical protocol e g 2 Step for 30 minutes to warm up the DNA Engine then measure the air temperature at the back air intake vents If more than one machine is involved measure the air temperature for each If the air intake temperature of any machine is warmer than 31 C use table 3 1 to troubleshoot the problem Some experimentation may be required to determine the best solution when more than one cause is involved After taking steps to solve the problem verify that the temperature of the air entering the air intake vents has been low ered using the procedure outlined above Table 3 1 Troubleshooting Air Supply Problems Cause Air circulation is poor Ambient air temperature is high Machine is in warm part of room Machines
52. chines and different protocols can be run independently on them Networked machines can help save time for a busy laboratory For ex ample in a DNA Engine controlled network protocols can be entered into the controlling machine and then be run on all the other machines connected to the network Networking Machines Without a Computer 9 2 Connecting the Machines Connect DNA Engines and DNA Engine Tetrads via the IEEE 488 port on the back of each machine see figs 2 3 and 12 4 using high quality cables Although several connection configurations are possible we recommend a simple daisy chain or star pattern The DNA Engine s IEEE 488 interface is fully compatible with both the IEEE 488 2 and the IEEE 488 1 standards Assigning Address Numbers to Machines Each machine must have a unique address number which is assigned from its control panel Address numbers cannot be assigned from the controller To assign an address number to a machine select Setup from the Main Menu then press Proceed The Setup Menu will be displayed Setup _REMOTE Printer Lid Version Select Remote then press Proceed A screen allowing choice of a port will be displayed Remote _SERIAL TEEE 488 Networking Mi Select IEEE 488 then press Proceed The remote addressing screen will be displayed Remote Serial _TEEE 488 Address 0 30 Q Enter an address number for the machine Set the controller DNA Engine s
53. chosen specify the printer s baud rate as for Choosing a Remote Port p 8 7 Using the Utilities Mi Choosing a Minimum Block Temperature for the Hot Bonnet To set a minimum block temperature below which the Hot Bonnet will automatically turn off select Lid from the Setup Menu then press Pro ceed The following screen will be displayed Lid Mode _MINIMUM Select MINIMUM and press Proceed The folowing screen will be displayed Lid Minimum Turn off heated Lid below 23 C Use the keypad to enter the temperature then press Proceed to enter the value and return to the main menu Choosing a Temperature Control Mode for the Hot Bonnet Two temperature control modes are available for the Hot Bonnet e Tracking Offsets the temperature of the heated inner lid at least a specified number of degrees Celsius in comparison to the tempera ture of the sample block e Constant Keeps the inner lid at a specified temperature C The DNA Engine is set for the tracking mode at the factory using an offset of 5 C which should be adequate for most reactions The constant mode is provided for unusual reactions requiring the inner lid to be hot at all times When using the constant mode specify a lid temperature at least 10 C higher than any temperature used in the protocol we suggest 105 C To choose a control mode select Lid from the Setup Menu then press Proceed The following scree
54. contents of lt MAIN gt Edit lt MAIN gt _CUSTOM1 CUSTOM2 CUSTOM3 QUIKSTEP 2 STEP 3 STEP Select the program to be edited then press Proceed If programs have been stored in custom folders The screen will list all the folders residing in the machine Edit _ lt MAIN gt lt FOLDER1 gt lt FOLDER2 gt Select the folder containing the program then press Proceed A list of all programs in the folder will be displayed Select the program to be edited CUSTOM1 is used in the following and all succeeding ex amples in this chapter then press Proceed In either instance after you press Proceed the first editing screen will be displayed see next section 7 2 Editing Programs W Y Tip To retain the original version of a program copy the program see Copying a Program chapter 8 and then edit the copy Editing the Program The first editing screen displays all three temperature control methods The program s current temperature control method is displayed in all capital letters Edit CUSTOM1 Control method Block Probe _CALCULATED Select a different temperature control method if desired then press Pro ceed The first three lines of the program will be displayed Edit CUSTOM1 92 0 for 0 10 63 0 for 1 00 3 GoTo 1 24 times Use the Select keys to scroll up and down through the program The cursor will progressively move to the step number and the
55. ctors on the back of the next Remote Dock in the series and so on fig 13 4 e When the RAD system has been completely set up connect the fan power supply s wall plug to a power source The remote system is transparent to the base unit i e the Dock Connec tor allows the base to control the Alpha unit in the Remote Dock as if it were in the standard configuration IMPORTANT Turn the base unit s power off when changing the type of sample block you are using Turning the power off resets the base allow ing it to recognize the new block If not reset the base unit assumes that the previous type of block is installed resulting in error messages and procedural faults The Remote Alpha Dock System M Figure 13 5 Flush mounting template 23 70 10 9 65 rly 2 84 74 Q a 168 20 16 6 D 165 00 Oxa D25 00 TYPICAL 1 01 DIMENSIONS IN MILLIMETERS AND IN NCHES de 13 7 Appendix A Safety Warnings and Guidelines Warning 4 Warning Warning 4 Warning 4 Caution Operating the PTC 200 DNA Engine or PTC 225 DNA Engine Tetrad before reading this manual can constitute a personal injury hazard Only qualified laboratory personnel trained in the safe use of electrical equipment should op erate these machines Do not open or attempt to repair the PTC 200 DNA Engine or PTC 225 DNA Engine Tetrad base the PTC 225 DNA Engine Tetrad power supply any Alpha unit or any accessory to the PTC
56. dels 2 4 opening 4 5 removing 4 5 Baud rate setting 8 8 Beep option See Programs options types of Bleach using in block 10 2 C Chill out liquid wax 4 7 Cleaning air vents 10 2 and biohazardous materials 10 3 and radioactive materials 10 3 4 16 chassis and block 4 16 removing oil from block 10 2 solutions to use 4 16 Condensation in tubes following holds 4 8 Control panel 2 2 keys 4 3 lights 4 3 using 4 3 D DNA Engine running protocols separately by block 12 7 simultaneosly on all blocks 12 8 DNA Engine Driver Software 9 4 DNA Engine Tetrad air supply requirements 12 6 Alpha units operating 12 7 cancelling protocol runs 12 8 cleaning 12 10 connecting power cables 12 5 environmental requirements 12 6 fuses changing 12 10 general information 12 2 installing 12 5 12 6 instant incubations running 12 8 layout back view 12 4 bottom view 12 4 control panel 12 3 front view 12 3 networking 12 9 packing checklist 12 5 power supply requirements 12 6 programming 12 9 robotics installations 12 6 specifications 12 2 troubleshooting 12 10 turning machine on 12 7 utilities for 12 9 Documentation conventions graphic iv terminology iv typographic iv Electromagnetic interference A 2 Environmental requirements 3 2 3 3 Error messages 11 2 Extend option See Programs options types of In 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual F FCC warning A 2 Fuses changing 10 3 H Hot Bonnet
57. during preheat so Alpha unit has been shut down Machines are not getting enough air or air being taken in is warmer than 31 C Machine is running a protocol consisting of many cycles of only a few seconds each Alpha unit heat sink does not have time to dissi pate heat generated by rapid cycl ing Eventually its maximum al lowable temperature is exceeded and Alpha unit is shut down Sensor malfunction has allowed base to heat block over its maxi mum allowable temperature 107 500 triggering automatic Alpha unit shutdown See causes for HS Overheated Program Terminated Heat sink sensor in Alpha unit and power supply sensor in base are not working so Alpha unit has been shut down Troubleshooting Mi Action Change to Calc control mode Alpha unit and or base needs servicing Contact MJ Research or your distributor Use 96V Alpha module Heated lid or base needs servic ing Contact MJ RESEARCH or your local distributor Make sure machine gets enough air and that tempera ture of air being taken in is 31 C or cooler see p 3 3 Correct air supply problems and run protocol again If error message persists base may need servicing Contact MJ RESEARCH or your local distributor Contact MJ RESEARCH or your local distributor to discuss protocol Alpha unit and or base needs servicing Contact MJ RESEARCH or your local distributor See actions for HS Overheated Program Terminated
58. e blue thumbwheel all the way counter clockwise to completely raise the inner lid 3 Load either a microplate or at least eight individual tubes into the sample block The inner lid pivots around a central point so it is im portant to distribute individual tubes evenly Load at least four tubes in the center of the block and at least one tube in each of the four corners of the block If using a sealing film or mat apply it to the loaded microplate according to the manufacturer s directions 4 Close the Hot Bonnet by pressing down on the top lever Turn the thumbwheel clockwise to lower the Hot Bonnet s inner lid onto the loaded microplate tubes The thumbwheel turns easily at first since the inner lid has not yet come into contact with anything Stop turn ing the thumbwheel when you feel increased resistance which indi cates that the inner lid has touched the microplate tubes 5 Open the Hot Bonnet Turn the thumbwheel clockwise an extra half to three quarters of a turn to set an appropriate lid pressure Caution Do not turn the thumbwheel more than three quarters of a turn This can make it hard or impossible to close the lid and puts excessive strain on the latch holding the lid closed An extra half to three quarters of a turn ensures the correct pressure for most types of reaction vessels Some empirical testing may be required to determine the optimum pressure required for certain vessels Once this pressure has been d
59. ed just before the End step Cancelling Editing Changes To cancel all editing changes made to a program use the Select keys to move the cursor to any step number then press Cancel A cancellation confirmation screen will be displayed Edit CUSTOM1 Cancel changes _YES No Select Yes then press Proceed All editing changes will be abandoned and the Main Menu will be displayed 7 5 Using the Utilities Locating a Stored Program 8 2 File Utilities 8 2 Creating a Folder 8 3 Assigning a Password to a Folder 8 3 Deleting a Folder 8 4 Copying a Program 8 4 Renaming a Program 8 5 Moving a Program 8 5 Deleting a Program 8 5 List Utilities 8 6 Viewing a Program in the LCD Window 8 6 Printing a Program 8 7 Setup Utilities 8 7 Choosing a Remote Port 8 7 Choosing a Printer Port 8 8 Choosing a Minimum Block Temperature for the Hot Bonnet 8 9 Choosing a Temperature Control Method for the Hot Bonnet 8 9 Determining the Software Version Number 8 10 8 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual Locating a Stored Program Many of the DNA Engine s utilities require you to locate a program stored in the machine The actions necessary to do this depend on whether your programs have been stored in the lt MAIN gt folder or in custom folders e If all programs have been stored in lt MAIN gt The screen will list all the programs that lt MAIN gt contains Copy lt MAIN gt
60. een broken into and their associated versions L in the example MJ RESEARCH periodically updates the DNA Engine s software to incor porate new features or allow new Alpha units to be used Most upgrades are available free of charge for units under warranty and may be installed into a DNA Engine electronically from a desktop computer Contact your MJ RESEARCH sales representative or an authorized distributor for details Occasionally upgrades may require a hardware change These upgrades require return of the DNA Engine base to MJ REsEaRcH or an authorized distributor EZ Networking Overview of Networking 9 2 Networking Machines Without a Computer 9 2 Connecting the Machines 9 2 Assigning Address Numbers to Machines 9 2 Running a Protocol on Networked Machines 9 3 Networking Machines with a Computer 9 4 Creating a Computer Controlled IEEE 488 Network 9 5 Creating a Computer Controlled RS 232 Network 9 5 9 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual Overview of Networking Up to 15 machines consisting of any combination of DNA Engines and DNA Engine Tetrads may be networked with or without a computer as the controller When networking with a computer all the machines are controlled from the computer s keyboard When networking without a computer a specially designated DNA Engine controls the other machines in the network Any combination of single and dual block Alpha units may be loaded into networked ma
61. ence of spending less time running reactions shorter protocols also help preserve enzyme activ ity and minimize false priming Cycling denaturations run under calculated control are usually optimal at 5 seconds Annealing extension steps can also be shortened but the periods for these will be reaction specific Calculated control provides for shorter protocols in three ways 1 Brief and precise block temperature overshoots are used to bring samples to temperature rapidly 2 Incubation periods are timed according to how long the samples not the block reside at the target temperature 3 The machine automatically compensates for vessel type and reaction volume Block Control Block control provides less accurate control of sample temperatures than calculated control provides Under block control the temperature of samples always lags behind the temperature of the block The length of the time lag depends on the vessel type and sample volume but typically is between 10 and 30 seconds Block control is chiefly used to run proto cols developed for other thermal cyclers that use block control Creating Programs Mi Probe Control Probe control is available for unusual circumstances that may require it Ordinarily though it should be used with caution While the DNA En gine will have no trouble heating the probe to the target temperature if the probe is seated or prepared differently from the sample tubes actual sample temperatu
62. ent rules may apply in different countries Anyone who intends to use MJ Research equipment to do PCR is encouraged to contact Hoffmann La Roche for more information at one of the addresses below In the United States In other nations Kathe Kiehn Dr Agnieszka Junosza Jankowski Licensing Manager PCR Licensing Manager Roche Molecular Systems Inc F Hoffmann La Roche Ltd 1145 Atlantic Avenue Building 222 350 Alameda CA 94501 USA CH 4002 Basel Switzerland 510 814 2970 41 61 687 3031 Fax 510 814 2977 Fax 41 61 687 2113 Contents Documentation Conventions ecccccccccccccceccececccceccececcececceceescecsecececeececescesuscecsecussscessssessesacsess TU Part I The DNA Engine Tes TPO UICHON o 1 1 2 Layoutand Spe CHCA Ons illa 2 1 AS ral A ostia 3 1 BCC LAU O sate aa 4 1 DRUMMING roto COS in es 5 1 GO Create PRO PT ANS a a 6 1 de OUI TORT AIS nd 7 1 S Usine the Utiel loose Lisiil 8 1 NEON OF KIO drid iaa 9 1 LO Manten eii 10 1 GIR bi vey Cho rol 101 0 E ROPA _ ERE 5 eras ore nr pny tart nessa See err 11 1 Part II The DNA Engine Tetrad 12 Information Specific to the DNA Engine Tetrad 0 ee eeseeeeseeeeeeeseeeeseeeeeeneees 12 1 Part III Accessories 13 The Remote Alpha Dock System ssssssssssesesesssesesesesesesesesescecsesesessesesssesesosesesesesesese 13 1 Appendix A Safety Warnings and Guidelines oonocnccnocononnnnononononcnnonancnnncononanonncnnnonos A 1 Appendix B How a Peltier Heat Pump Works
63. ep number then press Proceed The Edit Menu will be displayed Enter CUSTOM1 2 92 0 for 0 40 _EDIT Insert Step Option Delete Select Option then press Proceed and follow the procedure for adding an option To abandon the Edit Menu and return to the list of program steps press Cancel e To enter Edit mode Move the cursor to the number of the step you wish to edit then press Proceed See chapter 7 for information on entering Edit mode If you do not wish to enter Edit mode press Can cel The screen will return to the list of program steps e To delete the program Position the cursor on any step number and press Cancel A cancellation confirmation screen will be displayed Enter CUSTOM1 Cancel program _YES NO Select Yes then press Proceed The program will be deleted and the Main Menu will be displayed Deleting a Program To delete a program after at least one complete step has been entered see Revising During Programming p 6 14 To delete a program before one complete step has been entered repeat edly press Cancel The cursor will move backward through the step s 6 19 MW DNA Engine amp DNA Engine Tetrad Operations Manual values deleting them as it goes When all values have been deleted keep pressing Cancel to move backward through Enter mode screens until the Main Menu reappears at which point you can begin a new program Keeping a Permanent
64. erials Out of the Block 10 3 Changing the Fuses 10 3 10 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual Cleaning the DNA Engine 10 2 Cleaning the Chassis and Block Clean the outside of the DNA Engine and Alpha unit with a damp soft cloth or tissue whenever something has been spilled on it or the chassis is dusty A mild soap solution may be used if needed Clean block wells with swabs moistened with water 95 ethanol or a 1 100 dilution of bleach in water see the Twin Tower Block Operations Manual for instructions on cleaning the Twin Tower s slide slots If using bleach swab wells with water afterward to remove all traces of bleach Clean spilled liquids out of the block as soon as possible dried fluids can be difficult to remove Do not clean the block with caustic or strongly alkaline solutions e g strong soaps ammonia bleach at a higher con centration than specified above These will damage the block s protec tive anodized coating possibly causing electrical shorting If you use oil in the block a practice not recommended by MJ RESEARCH see Using Oil to Thermally Couple Sample Vessels to the Block chap ter 4 clean the wells whenever the oil has become discolored or contains particulate matter Use a swab to determine whether cleaning is needed Clean the block with 95 ethanol as described above Oil buildup must be prevented Old oil harbors dirt which interferes with vessel seating and
65. etermined the thumbwheel position that deliv ers it may be marked with a colored marking pen or piece of tape Note As an aid in gauging how much the thumbwheel has been turned mark it at the quarter turn positions or every sixth bump on the thumbwheel there are 24 total bumps 6 Close the Hot Bonnet Loading Sample Vessels into the Block When using a small number of tubes they should all be placed in the center of the block to ensure uniform thermal cycling of all samples Also load at least one empty tube in each corner of the block to ensure that the Hot Bonnet exerts even pressure on the sample tubes see Adjusting the 4 9 MW DNA Engine amp DNA Engine Tetrad Operations Manual Hot Bonnet s Lid Pressure p 4 8 To ensure uniform heating and cooling of samples sample vessels must be in complete contact with the block Adequate contact is ensured by always doing the following e Ensure that the block is clean before loading samples see chapter 10 for cleaning instructions e Firmly press individual tubes or the microplate into the block wells Using Oil to Thermally Couple Sample Vessels to the Block With two exceptions see below MJ RESEARCH does not recommend using oil to thermally couple sample vessels to the block for the fol lowing reasons e Calculated control protocols do not run accurately when oil is used e Oil traps dirt which interferes with thermal contact between ves
66. f heat ing and cooling is entered the maximum rate will be used Press Proceed The ramp rate will move to the second line of the screen and a line allowing entry of a finish temperature for the ramp step will be displayed Enter CUSTOM1 3 2 0 s Finish temp Enter the finish temperature for the ramp step 69 C is used in the ex ample below Enter CUSTOM1 3 2 0 s Finish temp 69 Press Proceed The finish temperature will be moved to the second line of the screen and a confirmation menu will be displayed on the last line Creating Programs W Enter CUSTOM1 3 2 0 s to 69 0 OK _YES No Option Select one of the displayed choices then press Proceed e Yes accepts the step and displays the Enter Menu again Use the En ter Menu to enter the next step in the program e No allows reentry of the ramp rate and finish temperature e Option displays the Options Menu see Modifying a Program Step with the Options p 6 11 Y Tip A ramp step will not hold the block at its finish tempera ture Instead as soon as finish temperature is reached the next step of the program will immediately be executed To hold the block for some period of time at a ramp step s fin ish temperature make sure a temperature step immediately follows the ramp step and program the hold at the finish temperature into this step Entering a GoTo Step To enter a GoTo step select GoTo from the Enter Menu
67. f the machine is currently ramping the block s temperature see chapter 6 for information on ramping A confirmation screen will be displayed Run 2 STEP Go on to next step _YES No Select Yes then press Proceed The protocol will advance to its next step Pausing a Running Protocol Press Pause to temporarily stop a running program The timer in the lower right hand corner of the runtime screen will be replaced by the word Pause Run 2 STEP 1 92 for 0 05 Cycle 1 Calc 92 PAUSE The samples are held at the displayed temperature until either the Pause or the Proceed key is pressed which causes the protocol run to re sume A protocol cannot be paused before the target temperature for a given step has been reached If Pause is pressed before this point the block continues heating or cooling until the target is reached and then the pro tocol is paused Stopping a Running Protocol 5 8 Press either Stop or Cancel to stop a running protocol A cancella tion confirmation screen will be displayed Run 2 STEP Stop 2 STEP Yes _NO Running Protocols W Select Yes then press Proceed to cancel the protocol The total run time for the protocol will be displayed Run 2 STEP PROGRAM CANCELED Total time 1 15 Press Proceed to return to the Main Menu Note Turning off the machine does not stop a running protocol The DNA Engine will assume the prot
68. fferent con nector Use a one piece molded connector of the type specified above Air Supply Requirements The DNA Engine requires a constant supply of air that is 31 C or cooler in order to remove heat from the Alpha unit s heat sink Air is taken in from vents at the front back and bottom of the machine and exhausted from vents on both sides see figs 2 1 2 3 and 2 4 If the air supply is inadequate or too hot the machine can overheat causing performance problems soft ware error messages particularly HS Overheating and Slow Block Cy cling and even automatic shutdowns Special attention should be paid to airflow and air temperature in robotics installations of DNA Engines Ensuring an Adequate Air Supply e Do not block the air intake vents Position the DNA Engine at least 10cm from vertical surfaces and other thermal cyclers greater distances may be required see be low Do not put loose papers under the machine they can be sucked into the air intake vents on the bottom of the machine 3 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual Do not allow dust or debris to collect in the air intake vents The bottom air vents are particularly liable to collect dust and debris sometimes completely clogging up Check for dust and debris every few months and clean the intake vents as needed Remove light col lections of dust with a soft bristle brush or damp cloth Severe col lections of dust and de
69. fication addition or extension to this warranty except in the form of the extended warranty out lined below MJ Research Inc is not responsible for special incidental or consequential damages resulting from any breach of warranty or under any other legal theory including downtime lost samples or experiments lost reagents lost profits goodwill damage to or replacement of equipment prop erty and any costs of recovering or reproducing experimental results and data Exclusions This warranty applies only to machines sold in the U S A Under no circumstance will MJ RESEARCH ship a repaired or replaced machine or grant a refund of purchase price to a user in a nation in which there was an authorized MJ Resgarcu distributor at the time of purchase THIS WARRANTY IS NOT TRANSFERABLE FROM THE ORIGINAL PURCHASER TO A SUBSEQUENT OWNER FURTHERMORE THIS WARRANTY DOES NOT APPLY TO INSTRUMENTS USED OUTSIDE THE U S A EXCEPT WHEN EXPRESSLY AUTHORIZED IN WRITING BY MJ RESEARCH INC U S Domestic Extended Warranty Optional MJ RESEARCH Inc will offer to each ORIGINAL PURCHASER of an MJ Research BRAND thermal cycler the opportunity to purchase an extension of the warranty coverage explained above for an additional two years The coverage must be purchased through a purchase order received by MJ RESEARCH Inc within 30 days of receipt of the offer of extended warranty or the offer to renew the extended warranty These offers apply only to machi
70. g That Air Is Cool Enough 3 4 Requirements for Robotics Installations 3 5 3 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual Packing Checklist After unpacking the DNA Engine check to see that you have received the following e One DNA Engine base e One Alpha unit more if additional units were ordered e Two spare fuses e One power cord e One probe if purchased e PTC 200 DNA Engine s PTC 225 DNA Engine Tetrad Operations Manual this document e Product registration card US customers only e Extended warranty application US customers only e DNA Engine Driver remote software request form If any of these components are missing or damaged contact MJ RESEARCH or the authorized distributor from whom you purchased the DNA En gine to obtain a replacement Please save the original packing materials in case you need to return the DNA Engine for service See appendix E for shipping instructions Setting Up the DNA Engine The DNA Engine requires only minimal assembly plugging in the power cord and inserting an Alpha unit Insert the power cord plug into its jack at the back of the machine see fig 2 3 for location of jack then plug the cord into an electrical outlet With the machine turned off insert an Alpha unit see Installing an Alpha Unit chapter 4 Caution Do not insert or remove an Alpha unit with the DNA Engine turned on electrical arcing can result Read the safety warning in appendix
71. gin to run Setting Up a Block Control Protocol A single screen asking about use of the heated lid will be displayed Run ICEBUKET Use heated lid Yes _NO 5 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual 5 4 Select Yes or No then press Proceed The protocol will begin running Setting Up a Calculated Control Protocol Three screens will be presented e Ascreen asking for sample vessel information Run 2 STEP Vessel Type _TUBES Plate Select from the options shown may include tubes plates and slides then press Proceed Choose Plate for polycarbonate microplates and Tube for polypropylene tubes or polypropylene microplates For blocks that hold only tubes choose Thin or Thick depending on the type of tube loaded Note See Using Calculated Control in the Twin Tower Block Opera tions Manual for information on specifying slide format e A screen asking for the sample reaction volume last line in example below Run 2 STEP Vessel Type TUBES Volume ul 10 Use the keypad to enter a sample volume in microliters then press Proceed If sample vessels are sealed with oil or wax include the volume of the oil or wax in the total sample volume entered Note Specify 10ul for any volume less than 10ul Note Verify that your vessel sealing system is adequately tight be fore performing 5 ul reactions e A screen asking about use of the heated lid Select Yes or
72. her option for the step Entering a Beep Option To enter a beep select Beep from the Options Menu for a temperature step or a ramp step then press Proceed The word Beep will be dis played on the third line of the screen and a confirmation menu will be displayed on the last line Enter CUSTOM1 1 92 0 for 0 30 Beep OK _YES No Option Select one of the displayed choices then press Proceed e Yes accepts the Beep option and displays the Enter Menu again Use 6 17 MW DNA Engine amp DNA Engine Tetrad Operations Manual the Enter Menu to enter the next step in the program e No cancels the Beep option e Option displays the Options Menu again if a temperature step is being modified Use the Options Menu to enter another option for the step Option will not be displayed if a ramp step is being modi fied since Beep is the only option available for ramp steps Revising During Programming 6 18 To change values in a program you are entering follow the procedures described below This editing method should be used to change just a few values at a time To make many changes or to delete or add entire steps use Edit mode see chapter 7 To Change the Last Value Entered or Menu Option Chosen Press Cancel The choice just made will be cancelled so that another value may be entered or another menu option chosen Press Proceed after changing a value so that the program will accept it To
73. hosen 12 7 MW DNA Engine amp DNA Engine Tetrad Operations Manual 12 8 When the desired block has been selected press Proceed to run the protocol Note To run different protocols concurrently press Block to return to the main screen and select a new protocol and a new block as de scribed in this section Any protocols already initiated will continue to run Running the Same Protocol on All Blocks If all the Alpha units installed in the DNA Engine Tetrad have the same type of block i e each has the same number of wells or four Twin Tow ers are loaded and no protocols are currently running on any of them the same protocol including an instant incubation may be run on all of the blocks at once To do this select a protocol or press Instant to run an instant incuba tion then press Block until All appears in the upper right hand corner of the displayed screen Run 2 STEP on ALL Block 96V The status indicator lights for all of the sample blocks will be flashing green Press Proceed to run the protocol on all the blocks simultaneously Running an Instant Incubation Instant incubations are run in the same way as for the DNA Engine see chapter 5 The incubation is initially assigned to the first available block in numerical order of quadrants press Block to assign it to a different block or to all of them if the same type block is present in all four quadrants Cancelling Pro
74. ies the desired block 22 in the example below Run 2 STEP on 22 Block Remote 60V 9 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual The machine s address number will be identified in the upper right hand corner of the screen Remote indicates that this is the network screen for a slave unit and 60V indicates the type of Alpha unit that has been selected The status light for the selected block will flash on the control panel of the machine holding the block If the the right or left block of a dual block Alpha unit has been selected an A or a B will appear after the address number e g 7 3A for the right hand block in a dual block Alpha unit loaded into quadrant 3 of aDNA Engine Tetrad at address 7 See figure 12 1 for a diagram of the layout of quadrants in the Tetrad Press Cancel at any time to abandon the selection process and display the Main Menu again When the desired block is identified in the LCD window press Proceed When you have worked through the screens that set up the protocol s tem perature control method the protocol will begin running The protocol s runtime screen will appear on the controller s LCD win dow The LCD window of the machine holding the active block will con tinue to show whatever screen it was displaying before the protocol be gan running Press Block to display the Main Menu on the controller again Repeat the procedure to run protocols o
75. individual values for each step To change an individual value in a step position the cursor on it and type the new value then press Proceed The new value will be dis played on the screen To cancel a change press Cancel The original value will be restored To add or delete a step or to modify a step with an option position the cursor on the step number then press Proceed The Edit Menu will be displayed for that step Edit CUSTOM1 2 63 0 for 1 00 _EDIT Insert Step Option Delete e Edit displays the previous screen again but with the cursor positioned on the first value for the step so that it may be changed e Insert allows a step to be added before the displayed step e Delete deletes the displayed step e Option allows an option to be added to the displayed step if it is a temperature or a ramp step 7 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual Inserting a New Step To insert a new step select Insert from the Edit Menu then press Pro ceed The Enter Menu will be displayed for the new step a new step 2 is added in the example below Edit CUSTOM1 Step 2 _TEMP GoTo Ramp End Use the Enter Menu to create the new step see Entering the Program s Steps chapter 6 When the step is complete select Yes from the confir mation menu then press Proceed The program being edited will be displayed again with the new step appearing among the listed steps
76. ing an uncompleted program 6 16 entering steps beep option 6 14 end step 6 11 extend option 6 13 GoTo step 6 10 Increment option 6 12 ramp step 6 8 temperature step 6 7 general process 6 5 Initiating a program 6 5 modifying block and probe control programs for ca 6 5 modifying programs designed for other ma chines 6 5 Naming a program 6 6 revising during 6 14 setting a lengthy hold time 6 7 Programs designing 6 2 6 5 choosing temperature control method See Temperature control methods translating protocol into programinto pro grams 6 2 using GoTo steps to shorten programs 6 3 editing 7 2 7 3 7 4 adding an option 7 4 cancelling editing changes 7 5 changing individual values in a step 7 3 changing temperature control method 7 3 deleting step 7 4 inserting new step 7 4 saving edited program 7 4 7 5 finding in machine 5 2 7 2 8 2 options options types of 6 2 6 12 program steps types of 6 2 renaming See Utilities file Protocols and power failure 5 9 factory installed F 1 factory installed 5 2 manually stepping through 5 8 pausing while running 5 8 printing logs for runs 5 7 5 8 sample log 5 7 running choosing block to run protocol on 5 3 choosing protocol to run 5 2 reading completion screen 5 6 reading runtime screen 5 5 5 6 setting up temperature control method 5 3 three steps of 5 2 stopping while running 5 9 protocols adjusting for sample vessel type 4 6 Remote Alpha Dock System air supply requirements 13
77. ion of resonance to the transformer design gives it extraordinary efficiency These design innovations has made the DNA Engine s power supply small in size universal in input and resis tant to noise C1 Appendix D Shipping Instructions for US Residents Users residing in the United States should follow these instructions for shipping a machine to MJ RESEARCH for factory repair or an upgrade Users outside of the United States should send ma chines to their distributor in accordance with shipping instructions obtained from the distributor 1 Call MJ Researcu to obtain a return materials authorization RMA number Machines returned without an RMA will be refused by the Receiving Department Thoroughly clean the machine removing excess oil and radioactive and other biohazardous substances To protect the health of our employees MJ ReseEarcH will not repair or upgrade any machine that is excessively oily or that emits ionizing radiation upon arrival at our fac tory PLEASE ELIMINATE ALL BIOHAZARDS Pack the machine in its original packaging If this has been misplaced or discarded call MJ RESEARCH to request shipment of packaging materials You can also request a loaner machine which will be provided if available a rental fee may apply You can use the loaner s packag ing to return the machine needing repair Remove the Alpha unit from the DNA Engine or DNA Engine Tetrad base before shipping All warranties are voided if a
78. layed Enter CUSTOM1 4 End OK _YES No This screen automatically enters End on on the second line of the screen next to a step number and displays a confirmation menu for the step on the last line of the screen Creating Programs Mi Select one of the displayed choices then press Proceed e Yes accepts the step stores the program and displays the Main Menu e No displays the Enter Menu so that additional steps can be added If you have created custom folders for your programs see chapter 8 choosing Yes brings up a screen listing the folders Save program 1n lt MAIN gt lt FOLDER1 gt lt FOLDER2 gt Select the folder you want to store the program in then press Proceed The program will be stored in the folder and the Main Menu will be displayed Modifying a Program Step with the Options The Options Menu is accessible from the confirmation menus of tempera ture steps and ramp steps To access the Options Menu select Option from the confirmation menu of a temperature or ramp step then press Proceed The Options Menu will be displayed on the bottom line of the screen Enter CUSTOM1 1 92 0 for 0 30 Option _INC Ext Beep e Inc modifies a temperature step with an increment option An incre ment option allows a progressive increase or decrease of temperature each time the step is executed in a GoTo cycle e Ext modifies a temperature step with an extend option An extend op tion
79. layed Remote Serial _TEEE 488 8 7 MW DNA Engine amp DNA Engine Tetrad Operations Manual 8 8 The cursor will be positioned at the port currently in use To choose the serial port select Serial then press Proceed The cursor will move to Serial and a line allowing a baud rate to be specified will be displayed on the last line of the screen Printer _SERIAL TEEE 488 Baud 9600 2400_1200 Select a baud rate then press Proceed The remote port assignment will change to serial and the Main Menu will be displayed To choose the IEEE 488 port select JEEE 488 then press Proceed The cursor will move to IEEE 488 and a line allowing the DNA En gine to be assigned an IEEE 488 address will be displayed on the last line of the screen Printer Serial _TEEE 488 Address 0 30 Q The machine s current address will be displayed To change it enter a number from 0 30 then press Proceed The remote port assign ment will change to IEEE 488 the address number will be assigned to the machine and the Main Menu will be displayed Note Do not choose 0 for the machine address unless the DNA Engine will be a controller in a network see chapter 9 Choosing a Printer Port To choose a printer port select Printer from the Setup Menu then press Proceed The port selection screen will be displayed Printer Parallel _SERIAL Select the desired port then press Proceed If serial was
80. le and the amount of heat pumped is changed by changing the amount of current passed Both direction and amount of current flow are dictated by a microprocessor in the Alpha unit allowing precise control of thermal cycling in the Alpha unit block B 1 Appendix C How a Switching Power Supply Works Almost all solid state electronic devices including the DNA Engine s thermoelectric module re quire direct current DC for operation However electric utilities supply low Hertz alternating current AC which varies in voltage and frequency from nation to nation The DNA Engine uses switching transistors combined with high frequency resonant transformers to convert the in coming AC to DC The power supply first chops the AC power into small bursts of energy over 100 000 per second with the aid of high current switching transistors called MOSFETs metal oxide semiconductor field effect transistors The energy bursts are channeled into a high frequency transformer By changing the duration of the bursts that charge the transformer s magnetic core pulse width modulation a specific voltage output can be maintained even when the incoming voltage varies between 100 and 240 volts in the case of the DNA Engine Because the incoming power is being chopped so rapidly the incoming frequency is unimportant it can even be DC Spikes and surges in the incoming power no longer pose a problem since they are chopped nearly to oblivion The addit
81. le blocks the Alpha unit family that accom modate many types of tubes microplates and slides Hot Bonnet heated lid or its remote controlled version the Power Bon net for oil free cycling Intuitive software with easy to read interface for quick and painless programming editing file management password protection and much more Choice of calculated sample temperature control for highest speed and accuracy or block or probe control for compatibility with proto cols designed for a variety of instrument types Space saving design for easy setup and transportation Instant Incubate feature for continuous temperature incubations Networking for up to 15 machines for convenient remote operation and documentation of runs Customizable factory installed protocols When multiple users or high throughput operations must be accommo dated the PTC 225 DNA Engine Tetrad is available Essentially consist ing of four DNA Engines placed in one chassis the DNA Engine Tetrad has the same features and ease of use of the stand alone DNA Engine Using This Manual This manual contains all the information you need to operate your DNA Engine or DNA Engine Tetrad safely and productively Chapter 2 acquaints you with the physical characteristics of the DNA Engine Chapters 3 5 present the basics of installing and operating the DNA Engine Chapters 6 and 7 describe programming the DNA Engine Chapter 8 outlines the utilities available for
82. ls block temperature to keep the probe vessel at the programmed temperature using feedback information from the thermistor See chapters 5 and 6 for information on programming protocols for probe control Note Because the thermal characteristics of a probe can never precisely match those of a sample calculated control is often a better choice than probe control Customizing the Probe Vessel For the most precise control of sample temperatures install the probe s thermistor in the same type of tube that the samples will be placed in This is particularly important when the sample tubes have much thicker walls than the probe s tube Follow these steps to customize the probe vessel 1 Cut the hinge to the probe tube s lid if there is one Remove the lid and the attached the amber colored thermistor 2 Remove the lid from the new probe tube Add oil to the probe tube as described below under Adding the Oil 3 Gently place the thermistor in the new tube and snap the lid closed Make sure that the lid from the original probe tube fits the new tube tightly The probe wire may touch the sides of the tube The ther mistor should rest on the bottom of the tube Caution The thermistor is extremely fragile Handle it with great care 4 11 MW DNA Engine amp DNA Engine Tetrad Operations Manual Adding the Oil Viscous oils are the best choice for the probe tube s representative sample They closely mimic the the
83. lus one volume of oil overlay Note Use these oils only in the proportions outlined above Using them in any other proportion for example 1 1 with sample tube volumes will lead to inaccurate sample heating 4 17 Running Protocols Running a Protocol 5 2 Choosing a Stored Protocol to Run 5 2 Choosing a Block to Run the Protocol On 5 3 Setting Up the Temperature Control Method 5 3 Setting Up a Block Control Protocol 5 3 Setting Up a Calculated Control Protocol 5 4 Setting Up a Probe Control Protocol 5 5 Reading the Runtime Screen 5 5 Switching Between the Runtime Screen and the Main Menu 5 6 Reading the Protocol Completion Screen 5 6 Printing a Log for a Running Program 5 7 Manually Stepping Through a Protocol 5 8 Pausing a Running Protocol 5 8 Stopping a Running Protocol 5 9 Resuming a Protocol after a Power Outage 5 9 Using the Instant Incubation Feature 5 10 5 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual Running a Protocol 5 2 Running a protocol on the DNA Engine involves three steps 1 Ze 3 Choosing a stored protocol to run Choosing a block to run it on if a dual block Alpha unit is installed Setting up the temperature control method Either a custom designed protocol or one of the factory installed resi dent protocols may be run See appendix F for descriptions of the resi dent protocols which may be edited to fit your needs see chapter 7 All the factory installed prot
84. machine is shipped with an Alpha unit installed If the Alpha unit also needs to be shipped pack it in its original packaging materials Write the RMA number on the outside of the box Ship the machine freight prepaid to the following address We recommend you purchase insurance from your shipper Ship to Repair Department MJ Research Inc 136 Coolidge Ave Watertown MA 02172 D 1 Appendix E Warranties U S Domestic Warranty Standard MJ Research Inc warrants NEW MJ RESEARCH BRAND THERMAL CYCLERS MODELS PTC 100 PTC 150 PTC 200 amp PTC 225 against defects in material and workmanship for a period of two years from the date of purchase If a defect is discovered MJ Research Inc will at its option repair replace or refund the purchase price of the THERMAL CYCLER at no charge to the cus tomer provided the product is returned to MJ Research Inc within the warranty period In no event will MJ RESEARCH Inc be responsible for damage resulting from accident abuse misuses or inadequate packaging of returned goods ANY implied warranties including implied warranties of the merchantability and fitness for a particular purpose are limited in duration to TWO years from the date of original retail purchase of this product The warranty and remedies set forth above are exclusive and in lieu of all others oral or written expressed or implied No MJ RESEARCH dealer agent or employee is authorized to make any modi
85. maximum rate of heating or cooling GoTo step optional Causes the program to cycle back to an earlier step for a specified number of times up to 9 999 times End step mandatory Instructs the DNA Engine to shut down its heat pump because the program is complete Additional instructions termed options can be added to certain pro gram steps to modify their effects 1 Increment Modifies a temperature step to allow a progressive increase or decrease of temperature 0 1 10 0 C per cycle each time the step is executed in a cycle This is useful in touchdown programs when the annealing temperature of an oligonucleotide is not known Extend Modifies a temperature step to allow progressive lengthen ing or shortening of a temperature step hold by 1 60 sec cycle each time a step is executed in a cycle This is useful for accommodating an enzyme with diminishing activity Beep Modifies a temperature step or ramp step to make the machine beep when the target temperature is reached Creating Programs Mi Designing a New Program Translating a Protocol into a Program Until you are completely familiar with programming the DNA Engine you may find it helpful to first translate the protocol into DNA Engine program steps and options on paper Write down the protocol to be pro grammed one step per line Then write the type of program step that goes with the protocol steps at the end of each line If a protocol
86. mewhat perme able both to water and the S breakdown product This problem is exacerbated when polycarbon ate plates are held at high temperatures for long periods of time or when the plates are sealed for oil free thermal cycling 0 2 ml Polypropylene Tubes and 96 Well Polypropylene Microplates These tubes are manufactured with very thin walls to enhance thermal transfer The thin walls are somewhat fragile and can craze or develop small cracks when subject to mechanical stress Undamaged thin polypropylene tubes may also be somewhat permeable to the S break down product Either way there have been reports of S passing through the walls of 0 2 ml tubes of several different brands during thermal cycling No data are yet available on radio active contamination with polypropylene microplates 0 5 ml Polypropylene Tubes Contamination problems are rarer with this type of tube but instances have been reported The Solution 1 Substitute the low energy beta emitter P in cycle sequencing P nucleotides are not subject to the same kind of chemical breakdown as S nucleotides and they have not been associated with volatile breakdown products 2 If S must be used three things will help control contamination an oil overlay inside the tubes mineral oil in the thermal cycler outside the tubes and use of thick walled 0 5 ml tubes Always run S thermal cycling reactions in a fume hood and be aware that vessels may be contaminated on
87. mpera ture control method will be displayed Enter CUSTOM1 Control method Block Probe _CALCULATED Select a control method then press Proceed Entering the Program s Steps When a temperature control method has been chosen the Enter Menu will be displayed Creating Programs Mi Enter CUSTOM1 Step 1 _TEMP Gradient Ramp End Use this menu to enter each step of the program e Temp enters a temperature step e Gradient enters a Gradient step e GoTo enters a GoTo step note since a protocol can not begin with a GoTo step GoTo does not appear on the initial Enter Menu e Kamp enters a ramp step e End enters the End step Entering a Temperature Step To enter a temperature step select Temp from the Enter Menu then press Proceed The first Temp screen will be displayed Enter CUSTOM1 es Temp C The second line of this screen shows the number of the step being programmed 1 is used in the example above The last line of the screen allows a target temperature in degrees Celsius to be entered for the step Use the keyboard to enter any number between 5 0 and 105 0 as the target temperature 92 0 is used in the example below Enter CUSTOM1 Temp C 92 0 Press Proceed The target temperature will move to the second line of the screen and a line allowing a hold time to be entered for the step will be displayed Enter CUSTOM1 1 92 0 Time Enter the h
88. n If error message persists software upgrade may be needed or base may need servicing Contact MJ RESEARCH or your local distributor Alpha unit lid needs servicing soon Contact MJ RESEARCH or your local distributor Contact MJ RESEARCH or your local distributor 11 5 MW DNA Engine amp DNA Engine Tetrad Operations Manual Problems Related to Protocols Error Message Reaction is working but broad low mole cular weight band is seen in gels Reaction working but unexpected extra products or smear is seen 11 6 Following is a general description of some common problems related to the protocols and reaction components in sequencing and amplification applications For a more detailed discussion of protocols and reactions see Current Protocols in Molecular Biology F Ausubel et al eds John Wiley amp Sons specifically chapters 7 DNA sequencing 14 in situ hybridiza tion and immunology and 15 polymerase chain reaction Cause and Result Primer dimer material often produces a broad band in the lt 100bp region of gels Nonspecific hybridization occur ring during setup Reaction component concentration too high or too low Annealing temperature too low Protocol contains a wrong value Template not of sufficient purity Multiple templates or host DNA in sequencing reactions Action If obtaining appropriate reac tion product s no need to change anything Minimize primer
89. n screen is displayed Run 2 STEP PROGRAM COMPLETE Total time 50 31 Certain error messages may also be displayed in this screen see chap ter 11 Press Proceed to return to the Main Menu Running Protocols M Printing a Log for a Running Protocol If the DNA Engine is connected to a printer you can print a log for the run that includes e The DNA Engine s serial number and software version e The protocol s name e The temperature control method for the protocol e A list of the protocol s steps e The runtime thermal data To print a runtime protocol log first ensure that a printer is connected to the machine See Printing a Program chapter 8 for information about compatible printers Follow the instructions under Choosing a Printer Port chapter 8 to prepare the DNA Engine to communicate with your printer As long as a printer is connected to the machine the following screen will appear whenever a protocol is run Run 2 STEP Printer output _YES No To print the protocol as it runs select Yes then press Proceed Note If no printer is connected to the machine the screen allowing you to select printer output is not displayed at all 5 7 MW DNA Engine amp DNA Engine Tetrad Operations Manual Manually Stepping Through a Protocol Once a protocol is running pressing Proceed gives you the option of immediately advancing the protocol to the next programmed step even i
90. n a fuse blows the DNA Engine immediately shuts down and cannot be turned back on The machine records the event as a power loss so if a protocol is running when a fuse blows the ma chine will resume the run when the fuse is replaced and power restored see Resuming a Protocol after a Power Outage chapter 5 4 Warning TheDNA Engine incorporates neutral fusing which means that live power may still be available inside the unit even when a fuse has blown or been removed Never open the DNA Engine base You could receive a serious electrical shock Opening the base will also void your warranty 1 Disconnect the power cord from the back of the instrument Move the power switch to the 0 off position 2 Insert one corner of a small flat head screwdriver just under the fuse plug fig 10 1A and gently pry the plug loose Pull the plug straight out as far as it will go then push it downward to expose the fuses fig 10 2B 3 Remove both fuses and replace them with new ones it is impossible to visu ally determine which fuse is blown You may also test the fuses with an ohmmeter to determine which is defective and replace just that one 4 Gently press the fuse cover back in place and reconnect the power cord Figure 10 1 A How to pull out the fuse plug B Location of the fuses in the opened plug Fuse plug Fuse Fuse plug Screwdriver 10 3 Troubleshooting Error Messages 11 2 Problems Related
91. n block screens and the Main Menu in the LCD win dow during a protocol run Status Indicator Lights e Power light Glows red when the DNA Engine is powered up e Block Status lights Indicate which blocks are in use glow red when blocks are heating and green when blocks are cooling Using the Data Ports The DNA Engine has three data ports located at the rear of the machine an RS 232 port an IEEE 488 port and a parallel printer port See chap ters 8 and 9 for information on using these ports to network machines connect them to a computer or print data 4 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual Operating Alpha Units Figure 4 1 Note Operation of the Twin Tower will not be discussed owing to the many differences between this type of Alpha unit and the others Please see the Twin Tower Block Operations Manual for operating instructions Note Alpha units equipped with Power Bonnets are installed and removed as described below See the Power Bonnet User s Manual for instructions on opening and closing Alpha units with Power Bonnets Installing an Alpha Unit 1 Turn the DNA Engine off see the Caution on p 4 5 2 Hold the Alpha unit at its front and back edges 3 Lower the Alpha unit into the DNA Engine base leaving at least 3cm between the front edge of the Alpha unit and the front of the base 4 Raise the handle at the back of the Alpha unit and slide the block forward as far as it will go
92. n other networked blocks Press Block on the controller to progressively display the runtime screens of all net worked blocks that are running protocols While protocols are running other DNA Engine functions Edit Enter Files etc can be accessed on any individual machine in the network from that machine s front panel Networking Machines with a Computer 9 4 Networked machines may be conveniently controlled by a computer us ing DNA Engine Driver software which allows you to e Read and run protocols on any networked machine e Monitor block temperatures on a real time graphical display e Save runtime data to the hard drive for later viewing in spreadsheet or graphical format e Verify results and keep records of runs Contact MJ RESEARCH or an authorized distributor if you do not already have a copy of this software A complete manual is provided with the software Also available from MJ RESEARCH is a list of ASCII commands used to communicate with DNA Engines and DNA Engine Tetrads which Networking Mi may be used to write your own driver software A computer controlled network can be connected via IEEE 488 cables or RS 232 serial cables In either case the computer acts as the controller of the network and uses the driver software to communicate with the networked machines Creating a Computer Controlled IEEE 488 Network To connect DNA Engines and DNA Engine Tetrads in a computer con trolled IEEE 488 netwo
93. n will appear Lid _MODE Minimum Select Mode then press Proceed to select a minimum block tem perature see Choosing a Minimum Block Temperature for the Hot Bon net above 8 9 MW DNA Engine amp DNA Engine Tetrad Operations Manual 8 10 The mode currently in use will be selected in the screen select a different control mode if desired then press Proceed to enter the mode and re turn to the main menu If the tracking mode is chosen a lid offset up to 45 C above the block but not to exceed 110 C must be specified A line for this will appear at the bottom of the screen Lid offset C 15 If the constant mode is cho sen a lid target temperature up to 110 C must be specified A line for this will appear at the bottom of the screen Lid target C _ In either case enter a number then press Proceed The tracking control mode will be implemented and the Main menu will be displayed Determining the Software Version Number At times it is necessary to determine the version number of the software installed in the DNA Engine e g to report a problem to MJ RESEARCH To do this select Version from the Setup Menu then press Proceed The current version number will be displayed Version 1 1L CTRL L EXEC L USER L COMM L The top line reports the software version number 1 1L in the example The other two lines refer to the four pages that the software has b
94. nes sold and used in the U S A E 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual Some states OF THE U S A do not allow the exclusion or limitation of incidental or consequential damages or limitations on how long an implied warranty lasts so the above limitation or exclu sion may not apply to you This warranty gives you specific legal rights You may also have other rights depending upon your state of residence All provisions of this warranty are voided if the product is resold repaired or modified by any one other than MJ RESEARCH or an authorized distributor E 2 Appendix F Appendix F Factory Installed Protocols Mi Factory Installed Protocols Note A programmed time of 0 00 00 means hold at this temperature forever Name ICEBUKET LIGATION 37 1 HR 37 6 HR 65 DENATURE BOIL CUT amp KILL Temp Control Method Block Block Block Block Block Block Block Block Temp amp Time 4 0 00 00 END 15 0 00 00 END 37 1 00 00 4 0 00 00 END 37 6 00 00 4 0 00 00 END 65 0 00 00 END 95 0 05 00 END 100 0 05 00 END 37 1 00 00 70 0 05 00 END F 1 Index A Accessories See Power Bonnet Air supply requirements ensuring adequate air supply 3 3 Ensuring air is cool enough 3 4 troubleshooting problems with 3 4 Alpha units closing 4 6 installing 4 4 maximum rate of heating and cooling 6 2 mo
95. ocol was stopped by a power outage and will resume running the protocol when the machine is turned back on see below Resuming a Protocol after a Power Outage If a power failure occurs when a protocol is running the DNA Engine will hold the protocol in memory for a minimum of 24 hours to a maxi mum of 7 days depending on environmental conditions When power is restored the protocol will begin running again at the point at which it was stopped and a notice about the power interruption will be displayed The notice will identify the step and the cycle that were running when the power failure occurred and the block s temperature at the time power was restored Run 2 STEP A C POWER FAILED Cycle 1 Step 1 Recovered at 20 2 Press Proceed to remove this screen The protocol s runtime screen will immediately be displayed 5 9 MW DNA Engine amp DNA Engine Tetrad Operations Manual Using the Instant Incubation Feature 5 10 The DNA Engine may be converted to a constant temperature incubator by pressing Instant Incubate A screen allowing use of the heated lid will be displayed INCUBATE Use heated lid Yes _No Use the Select keys to enable or disable the heated lid then press Pro ceed A screen allowing entry of the incubation temperature will be displayed INCUBATE 1 20 0 for ever Use heated Lid Yes _No Use the keypad to enter any incubation temperature from 5 0 C to
96. ocols are stored in a single folder called the lt MAIN gt folder at the time of shipping Choosing a Stored Protocol to Run With the Main Menu displayed select Run then press Proceed One of two types of screen will be displayed depending on whether custom protocols have been stored in the lt MAIN gt folder or in custom folders If all protocols have been stored in the lt MAIN gt folder The first of three or more screens listing the protocols will be displayed Custom protocols are listed first then the 14 factory installed programs Run lt MAIN gt _CUSTOM1 CUSTOM2 QUIKSTEP 2 STEP 3 STEP EXTEND Use the Select keys to scroll through the listed protocols Scroll past the last or first listed protocol to see the next screen down or up Select the desired protocol then press Proceed If custom protocols have been stored in one or more custom folders One or more screens listing all the folders residing in the machine will be displayed Run _ lt MAIN gt lt FOLDER1 gt lt FOLDER2 gt lt FOLDER3 gt Select the folder that contains the protocol then press Proceed One or more screens listing the protocols stored in the folder will be dis played Use the Select keys to scroll through the listed protocols Select the desired protocol then press Proceed Running Protocols W In either instance after you press Proceed a screen similar to the fol lowing example will be dis
97. ogram To rename a program select Rename from the File Menu then press Pro ceed Locate the program to be renamed see Locating a Stored Pro gram p 8 2 then press Proceed A naming screen will be displayed Rename CUSTOM1 Name the new program see Naming the Program chapter 6 then press Proceed The program will be renamed and stored and the Main Menu will be displayed Moving a Program To move a program select Move from the Files Menu then press Pro ceed Locate the program to be moved see Locating a Stored Pro gram p 8 2 then press Proceed A screen listing all folders will be displayed Select the folder the program should be moved to then press Proceed The program will be moved to the new folder and the Main Menu will be displayed Deleting a Program To delete a program select Delete from the Files Menu then press Pro ceed Locate the program to be deleted see Locating a Stored Program p 8 2 then press Proceed A confirmation screen will be displayed Delete CUSTOM3 Delete program _YES No 8 5 MW DNA Engine amp DNA Engine Tetrad Operations Manual List Utilities 8 6 Select Yes then press Proceed The program will be deleted and the Main Menu will be displayed To cancel the deletion press Cancel or select No and press Pro ceed Use these utilities available under List to accomplish the following ta
98. old time for the step 30 seconds is used in the example be 6 7 MW DNA Engine amp DNA Engine Tetrad Operations Manual 6 8 low Enter CUSTOM1 1 92 0 Time Note Ifa hold time of zero 0 is entered the DNA Engine will hold the block at the target temperature indefinitely Press Proceed The hold time will move to the second line of the screen and a confirmation menu will be displayed on the last line Enter CUSTOM1 1 92 0 for 0 30 OK _YES No Option Select one of the displayed choices then press Proceed e Yes accepts the step and displays the Enter Menu again Use the En ter Menu to enter the next step in the program e No allows reentry of the target temperature and hold time for the step e Option displays the Options Menu see Modifying a Program Step with the Options p 6 11 Y Tip Avoid programming many short holds of only a few seconds each This can overheat the block causing the HS Overheat ing or PS Overheating error messages to be displayed and triggering automatic Alpha unit shutdowns if the block exceeds its maximum allowable temperature Entering a Gradient Step When you reach the step at which a gradient is desired select Gradient from the Enter Menu and then press the Proceed key For the purpose of this example the step in which the gradient will be entered shall be Step 2 Enter CUSTOM1 Step 2 Temp Goto _GRADIENT Ramp End Crea
99. ons 6 16 Entering an Increment Option 6 15 Entering an Extend Option 6 16 Entering an Beep Option 6 18 Revising During Programming 6 18 To Change the Last Value Entered or Menu Option Chosen 6 19 To Change All the Values in the Step Being Entered 6 19 To Change Values for Earlier Steps in the Program 6 19 Deleting a Program 6 20 6 1 MW DNA Engine amp DNA Engine Tetrad Operations Manual The Elements of a Program 6 2 DNA Engine programs consist of a series of steps encoding a protocol These steps are run using one of three possible temperature control meth ods calculated control block control and probe control Programs may contain five types of steps Two of the steps are manda tory and two are optional 1 Temperature step mandatory Sets a temperature for the block and the length of time it is held at that temperature The DNA Engine brings the block to this temperature at its maximum rate of heating or cooling unless modifying instructions are added to the program The maximum rate of heating and cooling is up to 3 C sec for all single and dual block Alpha units 1 2 C sec for the Twin Tower Gradient step optional allows you to program a temperature gra dient across the sample block The range of any single gradient can be as great as 24 C from left to right The maximum programmable temperature is 105 C the minimum programmable temperature is 30 C Ramp step optional Sets a slower than
100. p also known as a thermoelectric module This solid state device is manufactured to withstand the thermal stresses associated with rapidly cycling temperatures A thermoelectric module consists of numerous pairs Figure A 1 A thermoelectric module of crystalline semiconductor blocks precisely sand wiched between two layers of ceramic substrate fig A 1 The blocks are of two varieties N type which AA has a surplus of electrons in its crystalline structure AS and P type which has a deficit of electrons The two ewer et ee eds types are positioned in alternating pairs within the in ar eerie nermost layer of the sandwich Metal conductor The two types of blocks are wired together in alternat ing pairs When electrical current is passed through the blocks electrons in the N type blocks and the holes or empty electron spaces in the P type blocks are ex cited at one conductor semiconductor interface which absorbs a small amount of heat The electrons and holes flow through the crystalline blocks and return to a low energy state at the other conductor semi conductor interface with the release of the previously absorbed heat A thermal gradient of up to 70 C can be generated across the blocks in this manner telluride P type bismuth 4 telluride EA y Power input The direction of heat pumping is reversed by reversing the polarity of current flow through the thermoelectric modu
101. p in the program e No allows reentry of the Inc value e Option displays the Options Menu again Use the Options Menu to enter another option for the step Entering an Extend Option To enter an extend option select Ext from the Options Menu of a tem perature step then press Proceed The first Ext screen will be dis played Enter CUSTOM1 1 92 0 for 0 30 Sec cycle Creating Programs Mi As for Inc the temperature step being modified appears on the second line of this screen The plus sign on the third line means that the screen is set up to enter progressive lengthening of hold time Press to switch to a minus sign allowing entry of a progressive shortening of hold time Press Cancel to change back to a plus sign Enter the numerical value of the increase or decrease in hold time 1 0 is used in the example below Enter CUSTOM1 1 92 0 for 0 30 Sec cycle 1 Press Proceed The Ext value just entered will move to the third line of the screen and a confirmation menu will be displayed on the last line Enter CUSTOM1 1 92 0 for 0 30 1 sec cycle OK _YES No Option Select one of the displayed choices then press Proceed e Yes accepts the Ext value and displays the Enter Menu again Use the Enter Menu to enter the next step in the program e No allows reentry of the Ext value e Option displays the Options Menu again Use the Options Menu to enter anot
102. pha Dock System M Packing checkist e One Dock Connector e One Remote Alpha Dock e One multi pin power cable e One multi pin data cable e One fan power supply RAD 0201 only e One wall plug power cord RAD 0201 only e Three round jack power cords RAD 0201 only e Product registration card US only e Extended warranty application US only Requirements Environment The Remote Alpha Dock System allows for custom installations The following placement configurations are recommended by MJ Research e Remote Docks with mounted Alpha units may be configured in any horizontal orientation or array as long as a minimum side clearance of 10 cm is maintained between the Remote Dock and any wall bulk head or adjacent Remote Dock unit this is identical to the PTC 200 PTC 225 base requirement Requirements for motorized lid opera tion or for loading or unloading plates may dictate additional clear ances Figure 13 2 Remote Alpha Dock and Dock Connector bottom view EES 13 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual Remote Docks with mounted Alpha units may be stacked vertically as long as a minimum bottom clearance is maintained that would be no less than that resulting from the unit being placed on a solid hori zontal platform A minimum top clearance is also required to allow access to and operation of the Alpha unit lid Remote Docks can be flush mounted i e with the feet removed
103. played Run 2 STEP Vessel Type _TUBES Plate The top line of the screen will identify the selected protocol 2 STEP in the example The other lines on the screen will request information needed to set up the temperature control method explained below Choosing a Block to Run the Protocol On If a dual block Alpha unit is installed or if networked Alpha units are available see chapter 9 one of the blocks must be designated to run the protocol The first available block is automatically designated when a protocol is chosen Press Block to choose a different block The selected block s letter is identified in the upper right hand corner of the screen Run 2 STEP on A Vessel Type _TUBES Plate The status indicator light for the selected block will flash green Setting Up the Temperature Control Method When the protocol is selected and the correct block is designated for a dual block Alpha unit one or more screens will be displayed These screens will ask for information needed to set up the block s temperature control method The DNA Engine can control the block s temperature in three different ways block control calculated control and probe con trol Chapter 6 explains in detail how these methods work and their im plications for protocol design The following describes how to respond to the screens that are displayed for each control method When these screens have been dealt with the protocol will be
104. re control method to calculated control Action Alpha unit needs servicing soon Contact MJ RESEARCH or your local distributor Same as previous Same as previous Fan must be repaired Contact MJ RESEARCH or your local distributor Alpha unit needs servicing soon Contact MJ RESEARCH or your local distributor Alpha unit needs servicing soon Contact MJ RESEARCH or your local distributor Base needs servicing Contact MJ ReseaRcH or your local distributor Delete unused protocols and folders from memory Reduce size of stored programs by using GoTo and the Inc and Ext options see chapter 6 Make sure probe is installed when running probe control programs If error message persists replace probe Error Message PS Overheated Program Terminated PS Overheating Check Air Flow PS Sensor Fault Temp Ignored Slow Block Cycling Service Alpha Soon Slow Lid Cycling Service Alpha Soon Unit Failure Unit X Failure Tetrad Cause and Result Machine is not getting enough air or air being taken in is warmer than 31 C Machine is running a protocol consisting of many cycles of only a few seconds each Alpha unit heat sink does not have time to dissipate heat generated by rapid cycling Eventually its maximum allowable temperature is exceed ed and Alpha unit is shut down See causes for PS Overheating Program Terminated Power supply sensor is not working properly Protocols can
105. res can vary widely from the probe s temperature Probe control also cannot be used with microplates or slides Modifying Block and Probe Control Programs for Calculated Control Probe control programs will generally run well under calculated control with no modification other than changing the method of temperature control Block control programs can be changed to calculated control by subtracting at least 15 20 seconds from each temperature step Some empirical testing may be required to adjust modified programs for opti mum performance Modifying a Program Designed for a Different Machine The ramp programming step can be used to adapt programs designed for thermal cyclers with slower maximum heating and cooling rates than the DNA Engine In addition a given protocol will occasionally work better with a slower rate of temperature change the ramp step can be used to optimize the program for such a protocol Entering a New Program Programming the DNA Engine moves through five steps 1 Initiating the program 2 Naming the program 3 Choosing a temperature control method 4 Entering the program s steps 5 Entering the End step Each step involves entering values from the keyboard or making selections from a menu Programs may be edited as they are being entered Programs are automatically saved when the End step is entered They are stored in the lt MAIN gt folder unless folders have been created for them 6 5 MW D
106. rk the computer must be equipped with an IEEE 488 GPIB interface card contact National Instruments MJ RESEARCH or an authorized distributor for information about purchasing this card Connect the machines and assign addresses as described under Net working Machines Without a Computer p 9 2 Make sure that none of the networked machines have been assigned IEEE 488 address 0 as this will cause communication problems with the computer Install the MJ RESEARCH DNA Engine Driver software on the computer and use it to control the network Creating a Computer Controlled RS 232 Network In a computer controlled RS 232 serial network each machine in the network is connected directly to the controlling computer via a serial port The number of machines that can be included in such a network thus depends on the number of serial ports available on the controlling computer Use high quality serial cables to connect the serial port on each machine see figs 2 3 and 12 4 to a serial port on the computer Use the Setup utilities to configure each machine for serial communications see Choosing a Remote Port chapter 8 and select the serial port and the 9600 baud rate Install the MJ Research DNA Engine Driver software on the computer and use it to control the network 9 5 110 Maintenance Cleaning the DNA Engine 10 2 Cleaning the Chassis and Block 10 2 Cleaning the Air Vents 10 2 Cleaning Radioactive or Biohazardous Mat
107. rmal characteristics of buffer solu tion which changes temperature sluggishly due to the high specific heat of water MJ RESEARCH recommends using heavy mineral oil for the following reasons e The calculations required to determine the correct volume of oil are easy e Itis widely available and inexpensive Add mineral oil to the probe tube in the following proportions 1 x the volume of the buffer in an individual tube plus 1 x the volume of oil overlay if one is used It is important to use the correct amount of oil so that the representative sample changes temperature at the same rate as the actual samples To add the oil open the sample tube and pipette in the appropriate amount The oil must completely cover the thermistor Light and heavy silicone oil may also be used but necessitate more com plex calculations to determine the amount to add to the probe tube See chapter appendix 4 C for information on using these oils Note Use only mineral oil or silicone oil as the representative sample Figure 4 4 Correctly inserted probe 4 12 Probe wire exiting to left of plug Probe plug seated in jack Operation W Do not use paraffin wax or Chill out liquid wax or the probe readings will not be accurate I Caution Do not use water saline or any other aqueous solution as a representative sample Aqueous solutions will de stroy the thermistor Loading and Connecting the Probe Seat
108. rom the DNA Engine Tetrad s power supply fig 12 4 must be connected to the base and the power supply must be plugged into an electrical outlet 4 Warning Disconnect the power supply from the wall outlet before connecting or disconnecting the signal or power cable The power supply is capable of generating high current DC power High current circuits should never be con nected or disconnected when electrical power is present electrical arcs may result which can damage the power supply and pose a hazard of electrical shock 12 5 MW DNA Engine amp DNA Engine Tetrad Operations Manual 12 6 The signal and power cables terminate in twist lock plugs To screw a plug in first make sure the power supply is not plugged into an outlet Press the plug into the socket and twist it to the right A snap will be felt when the plug completely seats in the socket a faint click is also audible when the plug to the power cable fully seats The signal and power cables are joined at their power supply end by a single connector that plugs into the power supply This connector is housed in a protective metal cage screwed onto the back of the power supply Note Do not open the protective housing there are no user service able parts within Power Supply Requirements The DNA Engine Tetrad requires 10A 200 240VAC 50 60 Hz The unit is not designed to operate on the ordinary 100V 120V household power commonly supplied in North America and
109. sels and the block Caution Ifyou use oil in the block use only mineral oil Never use silicone oil It can damage the Alpha unit One exception to this recommendation involves the use of volatile radioac tive S nucleotides A small amount of oil in the block can help prevent escape of these compounds See chapter appendix 4 B for important infor mation regarding safe use of these compounds in polypropylene tubes and polypropylene and polycarbonate microplates A second exception involves the use of thick wall 0 5 ml tubes Certain brands of these tubes fit poorly in the block in which case oil may somewhat improve thermal contact When Figure 4 3 A Probe B Location of probe jack 4 10 Probe tube Probe plug Operation W ever possible use high quality thin wall tubes intended for thermal cycling see chapter appendix 4 A for a tube and plate selection chart Using the Optional Probe The probe consists of a precision thermistor mounted in a thin walled plastic tube A thin wire encased in a small plastic tube runs from the thermistor to the probe s plug which is inserted into a slot at the back of the Alpha unit fig 4 3 A small amount of oil is added to the probe tube to serve as the representative sample The tube is loaded into the block where it can serve as the control reference for any programmed target temperature between 0 and 100 C When a probe control protocol is run the DNA Engine contro
110. service amp support 888 652 9253 Gi Fo General fax 617 923 8080 Z A TR M Sr l l f a a a CL a wa D National Sales Office gt aF e gt 4 gt 987 Tahoe Blvd 106 a E a a Incline Village Nevada 89451 gf Allorders 888 729 2165 b x Of Re b M Se fg a SS T REQ a Web site www mir com Es a y US 4 e Distributors Worldwide a 4 gt a a A k AXA LIO XXX X AXA A e A e A e ye ad A 7 id A 7 a A 7 a A P a pa pa pa pa Mo O A Yow a a a a f a Q P gp y Ss PO w g Ny ao xf gt g Ny uE Po w gx
111. sk e View and print program steps Viewing a Program in the LCD Window To view a program in the LCD window select List from the Main Menu then press Proceed Locate the program to be viewed see Locating a Stored Program p 8 2 then press Proceed What happens next de pends on whether a printer is connected to the DNA Engine e Ifa printer is connected A screen will be displayed asking whether the program should be sent to the printer List CUSTOM1 Printer output Yes _NO Select No then press Proceed The program will be listed in the LCD window The first listed screen will show the program s control method Press Proceed to view subsequent screens listing each program step including any options Use the Select keys to scroll up or down through the steps one line at a time To return to the Main Menu press Cancel at any time or press either the right Se lect key or Proceed when the last step appears on the screen e Ifa printer is not connected The program will immediately be listed in the LCD window You will not be given the option of printing at all Note Listed programs cannot be edited To edit programs select Edit from the Main Menu see chapter 7 Setup Utilities Using the Utilities Mi Printing a Program First ensure that the DNA Engine is connected to a printer If no printer is connected you will not be given the option of printing at all The DNA
112. still be run but machine should be serviced soon too repair sensor Alpha unit block has not reached target temperature within expect ed time Unit will begin beeping and will continue to beep until target temperature is reached pro tocol is manually progressed to its next step see p 5 8 or protocol run is halted Problem often re sults from machine not getting enough air or taking air that is warmer than 31 C Alpha unit lid has not reached target temperature within expect ed time Unit will begin beeping and will continue to beep until target temperature is reached pro tocol is manually progressed to its next step see p 5 8 or protocol run is halted The self test has failed because of a problem with the base or the Alpha unit Troubleshooting Mi Action Make sure that machine is getting enough air and that temperature of air being taken in is 31 C or cooler see p 3 3 Correct any air supply problems and run protocol again If error message persists base may need servicing Con tact MJ RESEARCH or your local distributor Contact MJ RESEARCH or your local distributor to discuss protocol See actions for PS Overheating Program Terminated Base needs servicing Contact MJ RESEARCH or your local distributor Make sure machine is getting enough air and that tempera ture of air being taken in is 31 or cooler see p 3 3 Correct any air supply prob lems and run protocol agai
113. t 92 C for denaturation Check for appropriate anneal ing temperatures of primers using available computer pro erams or empirical testing Use List utility to check temperature control method for protocol change if needed Check screen for probe failure error message Probe may need servicing or replacing Call MJ RESEARCH or your local distributor Fill probe tube with correct amount of oil see p 4 11 Spike a complete reaction mix with a control template and primer set Use only high quality tubes plates that fit block snugly Ensure that wells are free of foreign materials that would interfere with tube plate seating 11 7 MW DNA Engine amp DNA Engine Tetrad Operations Manual Problems Related to Environmental Conditions Setup and Maintenance Problem Frequent shutdowns due to overheating Frequent Slow Block Cycling HS Overheat and HS Overheating error messages Dust and debris clog ging up air intake vents 11 8 Cause Machine is not receiving enough air Air flowing into intake vents is not lt 31 C Failure to regularly check for buildup Action Make sure air intake vents are not obstructed by dust debris or paper Remove light collec tions of dust and debris with damp cloth Vacuum out heavy collections Remove any papers placed under the machine Posi tion machine at least 10cm from vertical surfaces Check temperature of air enter ing
114. the probe tube in the center of the block fig 4 4 p 4 12 If oil is used to thermally couple samples to the block it must also be used on the probe tube but see Using Oil to Thermally Couple Sample Vessels to the Block p 4 9 Plug the probe into the jack at the back of the block so that the wire is to the left of the plug fig 4 4 Detecting a Faulty Probe If the DNA Engine detects that the probe is broken or missing when a protocol begins running the protocol s temperature control method is automatically switched from probe control to calculated control and the following message is immediately displayed Run CUSTOM1 Probe Sensor Failure Used Calc Control If the probe malfunctions during a protocol run the temperature control method is also switched to calculated control and the run time screen changes to say Calc instead of Probe When the run finishes the fol lowing message is displayed Run CUSTOM1 CALC control Probe not present 4 13 Appendix 4 A Tube Microplate and Sealing System Selection Chart Key Reaction vessel fits block without modification O Reaction vessel must be cut to fit this block MJ Research Thermal Reaction Vessels Sealing Options for Oil Free Cycling Cycler Blocks 96v 48 60 30 16 16 Description MJ Research Microseal Microseal Microseal 8 Strip 12 Strip SelfSeal Frame Seal ial ed Pe fracasar e Pe pepe fe gt poemo
115. the step s tar get temperature is reached a timer begins running in the lower right 5 5 MW DNA Engine amp DNA Engine Tetrad Operations Manual hand corner of the screen The first digit is the minutes elapsed the two digits after the colon are the seconds elapsed For protocols using calculated control or probe control press the left Se lect key to display the block s temperature on the third line of the screen Run 2 STEP 1 92 0 for 0 05 Block 62 1 Calc 68 0 This screen shows only as long as the left Select key is pressed The runtime screen returns when you stop pressing the key Press the right Select key to see a screen listing the cycle number if a GoTo step is executing time elapsed so far for the protocol run and estimated remaining time left in the run Run 2 STEP Cycle 1 Total time 0 20 Est remain 1 01 51 This screen is also displayed only as long as the key is pressed The runtime screen returns when you stop pressing the key Switching Between the Runtime Screen and the Main Menu Press Block to toggle between the runtime display and the Main Menu This allows you to edit a stored program enter a new one print a pro gram run another protocol on a dual block Alpha unit or networked DNA Engine or use the file utilities while the protocol runs Reading the Protocol Completion Screen 5 6 When the protocol run finishes a long beep sounds and a notificatio
116. ting Programs W The gradient screen will appear Enter CUSTOM1 Lower Temp C Enter the lower limit temperature for the purposes of this example 50 then press Proceed The upper temperature screen will appear Enter CUSTOM1 50 00 Upper Temp C Enter the upper temperature for the purposes of this example 70 then press Proceed Use integers only decimals are not accepted The next screen requires you to enter a hold time for the temperature gradient step Enter CUSTOM1 2 50 0 to 70 0 Time Enter the hold time in the form of min sec Press Proceed A confirma tion screen will appear Enter CUSTOM1 2 50 0 to 70 0 for 0 30 OK Yes No Option Select Yes to enter the step into memory and proceed to the next step select No to reject or edit the current step To preview select Option then Preview Preview gives the predicted temperatures for each of the wells along the sample block s long axis The lines do not all appear in the display use the Select keys to scroll up or down Gradient Preview Column 1 50 left Column 2 51 8 Column 3 53 6 6 9 MW DNA Engine amp DNA Engine Tetrad Operations Manual 6 10 Editing a gradient step To edit a gradient step select Edit from the Main Menu The program will be displayed as follows List CUSTOM1 1 92 0 for 0 20 2 50 to 70 0 for 0 30 Use the Select keys to
117. to oil This purified paraffinic oil solidifies at 14 C and is liquid at room temperature By programming a hold at low temperature the wax 4 7 MW DNA Engine amp DNA Engine Tetrad Operations Manual 4 8 can be solidified at the end of a run A pipette can then be used to pierce the wax in the tubes and remove the samples The wax is dyed red to assist in monitoring its use The dye has no adverse effects on fluorescent gel analysis of reaction products Sealing with the Hot Bonnet and Caps or Film Sheets The Hot Bonnet s heated inner lid maintains the air in the upper part of sample vessels at a higher temperature than the reaction mixture This prevents condensation of evaporated water vapor onto the vessel walls so that solution concentrations are unchanged by thermal cycling The Hot Bonnet also exerts pressure on the tops of vessels loaded into the block helping to maintain a vapor tight seal and to firmly seat tubes or the plate in the block Caps film or mats must be used along with the Hot Bonnet to prevent evaporative losses Tight fitting caps do the best job of preventing vapor loss and should be used for long term storage of reaction products Microseal A film is a quick way to seal Multiplate and Concord microplates or large arrays of tubes This film is specially designed to seal tightly during cycling yet release smoothly which minimizes the risk of aerosol formation and cross contamination of samples
118. tocols Press Block to select the block that is running the protocol to be cancelled then press Cancel A cancellation confirmation screen will be displayed Run 2 STEP on 2B STOP 2 STEP on 2B Yes _NO Information Specific to the DNA Engine Tetrad Mi Select Yes then press Proceed A screen announcing the cancellation and citing the total time for the run will be displayed Run 2 STEP on 2B PROGRAM CANCELED Total time 1 10 Programming the DNA Engine Tetrad The DNA Engine Tetrad is programmed in the same way as the DNA Engine see chapter 6 Networking the DNA Engine Tetrad Any combination of DNA Engine Tetrads and DNA Engines can be net worked as long as the 15 machine limit is not exceeded and a computer or a DNA Engine is included to serve as the controller the DNA Engine Tetrad cannot serve as controller See chapter 9 for information on net working machines Software Required for Networking ADNA Engine must have software version 1 1J or later in order to control a network that includes one or more DNA Engine Tetrads To identify the software version loaded into a DNA Engine see Determining the Software Version Number chapter 8 Operating Networked DNA Engine Tetrads Networked DNA Engine Tetrads operate the same as networked DNA Engines Each block of anetworked DNA Engine Tetrad is designated by the machine s IEEE 488 address followed by the block s quadrant num ber and an
119. ty 0 2m1 tubes in a number of styles including individual tubes and strips Microplates A variety of 96 well polycarbonate or polypropylene microplates can be used in 96V Alpha units as long as they fit the wells snugly Polypropy lene microplates are usually preferred because they exhibit very low pro tein binding and unlike polycarbonate microplates do not lose water vapor through the vessel walls This allows smaller sample volumes to be used as little as 5 10ul Polypropylene microplates and compatible Microseal A film mats or strip caps for sealing are available from MJ RESEARCH See Sealing with the Hot Bonnet and Caps or Film Sheets p Operation W X for a description of Microseal A Thin Walled Vs Thick Walled Tubes The thickness of sample tubes directly affects the speed of sample heat ing and thus the amount of time required for incubations Thick walled tubes delay sample heating since heat transfers more slowly through the tubes walls For the earliest types of thermal cyclers this delay mattered little These machines ramping rates were so slow below 1 C sec that there was plenty of time for heat to transfer through the tube wall to the sample during a given incubation Modern thermal cyclers have much faster ramping rates up to 2 3 C second so the faster heat transfer provided by thin walled tubes allows protocols to be significantly shortened For example in the reaction illus
120. ulated control The DNA Engine adjusts the block s tempera ture to maintain samples of a specific volume in a specific vessel type 6 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual 6 4 at programmed temperatures This includes optimized overshoots of the block by a few degrees for a few seconds which bring the samples to the programmed temperatures e Block control The DNA Engine adjusts the block s temperature to maintain the block at programmed temperatures independent of sample temperature e Probe control The DNA Engine adjusts the block s temperature to maintain the probe at programmed temperatures Calculated Control Calculated control is the method of choice for most types of programs yielding the most consistent most reliable and fastest programs When using calculated control the DNA Engine maintains a running estimate of sample temperatures based on the block s thermal profile the rate of heat transfer through the sample tube or slide and the sample volume or mass this information about the samples is provided when a program is run see Setting Up the Temperature Control Method chapter 5 Since this estimate is based on known quantities and the laws of thermody namics sample temperatures are controlled much more accurately than with block or probe control Hold times can be shortened significantly when protocols are run under calculated control In addition to the simple conveni
121. w Figure 12 4 Alpha units removed IEEE 488 port LY CD Electrical outlet plug Bottom View Figure 12 5 SS SSS GSS E E E MEU EU MEE O CD E E E E E O Y E E E E E E O CD E E E E E E E Ca Ca E CE gt C gt ll Jl Il Il ll Jl Il ll il il Information Specific to the DNA Engine Tetrad Mi Unpacking and Installing the DNA Engine Tetrad Packing Checklist After unpacking the DNA Engine Tetrad check to see that you have received the following e DNA Engine Tetrad base e Four Alpha units e Power supply unit e 220V power cord set inside power supply box e Four spare fuses e PTC 200 DNA Engine amp PTC 225 DNA Engine Operations Manual this document e Software request form US customers only e Extended warranty form US customers only e Warranty registration card US customers only If any of these components are missing or damaged contact MJ RESEARCH or the authorized distributor from whom you purchased the DNA En gine Tetrad to obtain a replacement Please save the original packing materials in case you need to return the Tetrad for service See appendix E for shipping instructions Installation The DNA Engine Tetrad requires minimal assembly installing the Alpha units see chapter 4 and connecting the signal and power cables to the back panel of the base Connecting the Cables The signal and power cables running f
122. worked together in any combination but a DNA Engine or a computer must serve as the controller unit for the network Networked DNA Engine Tetrads operate like networked DNA Engines with a few minor exceptions explained be low The Power Bonnet remote controlled heated lid may also be used with networked DNA Engine Tetrads if the controller is a computer The DNA Engine Tetrad s specifications are identical to those of the DNA Engine except for weight and size see chapter 2 Information Specific to the DNA Engine Tetrad Mi Layout Front View Figure 12 2 Dual block Alpha unit N NR AA E Power supply x JEN A AS gt o AP le o M A ogie Topog 9 henting Aunay Arena bike ni BS py Ment Ep lor Mes Ms DE 4 FIC225 Peltier Thermal Ochr Power supply air intake vent Front air exhaust vents Control panel Control Panel Figure 12 3 Instant Block status incubation lights key LCD display Cancel key Block Status DNA Engine Tetrad Y OOO OOO Instant OF y e Heating e Cooling CAUTION Sample blocks may be very hof ATTENTION Les blocs peuvent etre tres chduds C Select gt A DODO Block key Left and right selection keys Proceed key Single block Alpha unit Tetrad base Power light MJ RESEARCH KX PTC 225 Peltier Thermal Cycler O Power key 12 3 MW DNA Engine amp DNA Engine Tetrad Operations Manual Back Vie
123. y 90 noncondensing e Transient overvoltage per Installation Category II IEC 664 e Pollution degree 2 in accordance with IEC 664 Electromagnetic Interference The PTC 200 DNA Engine and PTC 225 DNA Engine Tetrad have been tested and found to com ply with the limits for a Class A digital device pursuant to part 15 of the US FCC Rules These limits are designed to provide a reasonable protection against harmful interference when the equip ment is operated in a commercial environment These machines generate use and can radiate radiofrequency energy and if not installed and used in accordance with the instruction manual may cause harmful interference to radio communications Operation of these machines in a resi dential area is likely to cause harmful interference in which case the user will be required to correct the interference at his or her own expense In addition the PTC 200 DNA Engine and PTC 225 DNA Engine Tetrad designs have been tested and found to comply with the EMC standards for emissions and susceptibility established by the European Union at time of manufacture FCC Warning Changes or modifications to the PTC 200 DNA Engine or PTC 225 DNA Engine Tetrad not ex pressly approved by the party responsible for compliance could void the user s authority to oper ate the equipment A 2 Appendix B How a Peltier Heat Pump Works The functional heart of every DNA Engine is a high performance Peltier effect heat pum

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