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S1000™ Thermal Cycler

1. 1000 Thermal Cycler Instruction Manual Catalog 184 2000 185 2096 185 2048 185 2384 Copyright 2007 Bio Rad Laboratories Inc Reproduction in any form either print or electronic is prohibited without written permission of Bio Rad Laboratories Inc Chill Out iProof iTaq and the helix logo are trademarks belonging to Bio Rad Laboratories Inc Other brands or product names are trademarks of their respective holders LICENSE NOTICE TO PURCHASER Purchase of this instrument Serial No conveys a limited non transferable immunity from suit for the purchaser s own internal research and development and for use in applied fields other than Human In Vitro Diagnostics under one or more of U S Patents Nos 5 656 493 5 333 675 5 475 610 claims 1 44 158 160 163 and 167 only and 6 703 236 claims 1 7 only or corresponding claims in their non U S counterparts owned by Applera Corporation No right is conveyed expressly by implication or by estoppel under any other patent claim such as claims to apparatus reagents kits or methods such as 5 nuclease methods Further information on purchasing licenses may be obtained by contacting the Director of Licensing Applied Biosystems 850 Lincoln Centre Drive Foster City California 94404 USA 1000 Thermal Cycler Manual Safety and Regulatory Compliance The S 1000 thermal cycler heats and cools quickly during operation We st
2. Create a two step protocol by combining annealing and extension into a single step Most polymerases remain active throughout the typical range of annealing temperatures 55 to 70 C Reduce the total run time by creating a two step protocol that combines the annealing and extension steps in a single step A two step protocol can produce a yield that is similar to a three step protocol for target sequence up to 200 bp To create a two step protocol keep the annealing temperature step and omit the extension step Then adjust the hold time for the annealing step based on the length of your amplicon Start with a hold time that is 10 seconds per 100 base pairs of the target Alternatively optimize the annealing temperature using a temperature gradient across the block Step 6 on page 23 and pick the final annealing temperature from the best results of the gradient experiment See Optimizing Temperature Steps With a Gradient page 55 for an example Optimization of the annealing step is critical because it determines the specificity of the reaction If the annealing temperature is too high the primers do not anneal easily and if the annealing temperature is too low the reaction will result in lower PCR yield primer mismatches and nonspecific amplification Optimize temperature steps to minimize the ramping time The larger the temperature difference between two successive steps in a protocol the longer the time required to reach
3. Protocol File Contents The protocol file contains both temperature steps and instrument settings that can be used to optimize a PCR reaction In the 1000 thermal cycler the temperature TEMP and gradient GRAD protocol steps create the temperature steps found in a typical PCR protocol However the GOTO and END protocol steps differ Read this section for detailed information about the content and structure of 1000 thermal cycler protocol files e Overview of the four types of protocol steps page 61 e Overview of instrument options that can be added to a protocol step page 64 Protocol Steps A PCR protocol includes temperature steps where the sample is kept at a target temperature for a specific amount of time called the hold time In the S1000 thermal cycler protocol file these temperature steps are called temperature TEMP and gradient GRAD steps A gradient step is a specialized temperature step that includes a temperature gradient across the block 61 1000 Thermal Cycler Manual Protocol Files and Folders 62 In general a PCR protocol includes these types of steps 1 An initial temperature step This initial temperature step denatures the template and also activates the polymerase To add an initial step in an S1000 thermal cycler protocol file add a temperature step page 22 Cycle of several temperature steps that amplify the PCR product These steps repeat in a cycle that amplifies the PCR pro
4. 00000 cece eee ene e ene 109 Components of the 1000 serieS 0000 e eee eee 109 Accessories for the 1000 SerleS 0 000 cee eee ees 111 1000 Thermal Cycler Manual Table of Contents 1000 Thermal Cycler Manual 1 Introduction Congratulations on purchasing the S1000 thermal cycler a member of the Bio Rad 1000 series thermal cycling platform It combines powerful thermal performance with unsurpassed flexibility and expansion potential making it ideal for both large and small laboratories This chapter provides an introduction to the S1000 thermal cycler and more including e The 1000 series platform of thermal cyclers below e 1000 thermal cycler features and performance page 2 e Bio Rad s PCR resources and support page 3 e Writing conventions for this manual page 4 The Bio Rad 1000 Series Platform The 1000 series line of PCR instrumentation is a flexible and modular platform designed to offer best in class performance and advanced features in an easy to use reliable package With the 1000 series platform you can get results faster by seamlessly integrating your workflows with functions such as PC compatibility optimization programming options online support and USB connectivity Table 4 1000 Series Catalog Numbers Product Catalog Number Item Description C1000 Thermal Cycler 184 1000 C1000 thermal cycler chassis only C1000 Fast 96 well System 185 1096 C1000 chassis Fast 96 well Rx
5. Adding an Increment INC to a Temperature Step The INC parameter changes the target temperature of a protocol step The increment can increase or decrease the target temperature with each cycle in the protocol To add an increment follow these instructions 1 Choose OPTION to add more parameters to a temperature step Press the right and left arrow keys to choose OPTION Then press ENTER to continue to the next screen Adding INC EXT RATE and BEEP Parameters In this example the OPTION will be added to step 3 parameters Step 3 n T oH oH Choose OPTION Tome T TO NOTE The INC parameter must be added to a step within the GOTO repeats in order to increment with each cycle of the reaction Choose INC to add an increment to the protocol step each cycle Press the right and left arrow keys to choose the INC parameter if it is not already selected Then press ENTER to continue to the next screen ay al ee ee Choose INC set FATE BEEP Enter the increment temperature Press the number keys to type an increment parameter To decrease the temperature each cycle enter a negative number by pressing the CANCEL key Press the PAUSE key to enter a decimal point Then press the ENTER key to continue to the next screen NOTE Enter an increment from 10 0 to 10 0 C per cycle in tenths of a degree and within the limits of a temperature step 0 100 C In this example the increment is 0 5
6. Amplifier temperature sensor 2 has failed Reaction module has been removed Protocol terminated Protocol storage memory has been corrupted Unable to read information from reaction module properly Information read from reaction module appears incorrect There was a problem with the block and power was shut off One of the block sensors in a single block has failed The system has cancelled the protocol at step x cycle y and sent the block to 4 C to preserve the samples The error message may instruct the user to contact Bio Rad In this event call the nearest Bio Rad Laboratories Technical Support team page 3 98 Sensor Failure means that the sensor was deemed short open or had changed more than 3 C in a 50ms period and that this condition was present for more than 2 seconds If two or more block sensors fail or both lid sensors fail the protocol is terminated and the block is sent to 4 C to preserve samples 99 1000 Thermal Cycler Manual Vaintenance amp Troubleshooting 100 1000 Thermal Cycler Manual Appendix A Preinstalled Protocols The S1000 comes with preinstalled protocols that provide a template for running new PCR experiments Use these protocols as templates The following sections describe the preinstalled protocols and include tips about when to use them Standard 2 and 3 step protocols below Touchdown PCR protocol to avoid non specific primer binding page 102 iTaq pr
7. Check the Firmware Version With the VERSION Tool When you want to check the firmware version that is currently on the thermal cycler select the TOOLS gt VERSION tool Follow these instructions to view the version 1 Select VERSION To select VERSION press the arrow keys Then press ENTER to continue to the next screen Select VERSION noc a ae ni a7 4 ae DE GR CO PO D 2 View the firmware version View the firmware version on this screen Additionally the header version of the alpha is also shown which means the firmware expects at least the specified version Press ENTER to return to the main menu WERSION HERDER Name the Thermal Cycler with the NAME Tool To name or rename the thermal cycler select the TOOLS gt NAME tool The S1000 thermal cycler name is displayed on the main menu Follow these instructions to name the thermal cycler or change the existing name 1 Select NAME To select the NAME press the arrow keys then press ENTER Select NAME 82 The TOOLS Options 2 Enter a name Enter a name by pressing the UP amp DOWN ARROWS to select a letter and pressing the number keys to type a number Then press ENTER to accept the selection and continue to the next character in the name NOTE A cycler name can contain from one to eight characters The characters can be numbers or capital letters In this example the letter M is entered by pressing the UP ARROW 12 times
8. Enter Cycler Hame Enter name The name or serial number identifies the S1000 thermal cycler when it is controlled by the C1000 thermal cycler page 88 The thermal cycler name appears in the C1000 thermal cycler instrument tree or in the 1000 series software instrument tree If the S1000 thermal cycler is not named then the S1000 thermal cycler serial number will be used 3 Confirm the name Press the ENTER key to confirm the thermal cycler name and to return to the main menu CYCLER HAME Enter Cycler Hamz Finish name MHEHDEL and press ENTER Change Default Parameters With the DEFAULTS Tool When you want to change the default parameters on the thermal cycler select TOOLS gt DEFAULTS These three parameters apply for a new protocol or instant incubation e Lid Target Sets the default lid temperature for new protocols page 33 e Turn off below Sets the temperature at which the lid heats to 31 C during a protocol run page 33 e Sample Vol Sets the default sample volume Vo1 in a new protocol Step 4 on page 21 Follow these instructions to view and change these default parameters 83 1000 Thermal Cycler Manual Advanced Tools and Functions 1 Select DEFAULTS Press the arrow keys to select DEFAULTS Then press ENTER to continue to the next screen Select DEFAULTS TO A TMM so mio H n a on rai a kee am gt T g oS a T m m eraa a T Ke TI y D Ole cet i wien a
9. Strip tubes low profile 0 2 ml 8 tube TLS Microplates Film MS Caps TCS Microplate Tube Sealer and Cap Supplies Table 10 Compatible sealing options for PCR microplates and tubes TUBES AND MICROPLATES CAPS AND SEALERS Unskirted and full height 24 48 and 96 well MLP Unskirted and low profile 48 and 96 well A B F P MLL Skirted and full height 96 and 384 well MSP Skirted full height and 96 and 384 well hard shell HSP The prefix of the catalog number for each series of supplies 59 1000 Thermal Cycler Manual Optimizing PCR With the S1000 Thermal Cycler 60 1000 Thermal Cycler Manual 6 Protocol Files and Folders This chapter describes the content and management of protocol files The VIEW and FILES options in the main menu provides quick access to the functions you need to view protocol file content and manage protocol files Read this chapter to learn about the following topics e Overview of the content of protocol files including step options and instrument settings page 61 e Overview of the management of protocol files and folders with the FILES option page 65 e Manage protocol files including how to copy move delete and rename files page 65 e Secure protocol files in a folder by entering a password page 71 e Manage folders including how to create delete secure and rename folders page 72 e View protocol files with the VIEW option page 75
10. 2 Change the default settings optional Press the up and down arrow keys to select a setting This example shows the system default parameters for the S1000 thermal cycler DEFAULT SETTING Lid temperature Block A Lid Target 14SM Default Lid turn off belo Turn off below ci BAE parameters temperature Sample Mol if pl NOTE The DEFAULT SETTINGS have the following ranges that apply to new protocols e Lid Target temperature setting can range from 0 to 110 C The default is set to 105 C e When the block is running an infinite hold at a temperature below the Turn off below parameter the lid heater maintains 31 C e Sample Vol setting can range from 0 to 50 uL or 30 uL for 384 well reaction modules and the default system parameter is 10 uL e Press the number keys to enter changes to the parameters Press ENTER to confirm the number and move to the next parameter 3 Save the changes Select YES or No Select YES and press ENTER to save the parameters Alternatively press the right arrow key to select No and press ENTER to reject changes and return to the main menu View a Temperature Gradient with the GRADCALC Tool If you want to review a temperature gradient to determine the temperature in each row select the TOOLS gt GRADCALC tool Select this tool to recreate the temperature in each row of wells across a block In the 1000 thermal cycler the gradient is distributed from the front Row H coolest tem
11. 10 0 Applies only to a temperature step increment to 10 0 C per cycle in page 28 Instructs the thermal cycler tenth of a degree to increment change the target temperature of a step with each cycle where a positive number increases the temperature and a negative number decreases the temperature EXT A time from 60 to 60 Applies to both temperature and extend seconds per cycle gradient steps page 30 Instructs the thermal cycler to extend the hold time with each cycle where a positive number increases the hold time and a negative number decreases the hold time RATE A number from 0 1 to 5 C Applies only to a temperature step thermal second page 31 Instructs the thermal cycler ramp rate to ramp to the target temperature at the specified ramp rate in that step Applies only to the Dual 48 and 96 Fast reaction modules BEEP No parameters Applies only to a temperature step page 32 Instructs the thermal cycler to beep as a signal that the thermal cycler reached the target temperature of that step PREVIEW No parameters Applies only to a gradient step Step 6 on page 23 Instructs the thermal cycler to displays the temperature gradient on the block Managing protocols With the FILES Option Managing protocols With the FILES Option The FILES option contains all the functions you need to manage protocol files and the folders that contain them Select the FILES option from the main menu to see these f
12. 1000 Thermal Cycler Manual 7 Advanced Tools and Functions The S1000 thermal cycler includes several advanced tools for stand alone and remote control The TOOLS option provides a list of tools including default settings and port settings Through USB or serial RS232 connections the 1000 thermal cycler can also be controlled externally by the C1000 thermal cycler or by a robotic system This chapter describes the tools listed in the TOOLS option and introduces the options for controlling the S1000 thermal cycler with another instrument e Select the TOOLS option to view or change instrument settings below e Control the 1000 thermal cycler with the C1000 thermal cycler and software through a USB port page 88 e Run the 81000 thermal cycler under automated robotic control through a USB or serial port page 90 The TOOLS Options The TOOLS option in the main menu provides a list of instrument settings and tools Select the TOOLS option in the main menu to see the list of tools The TOOLS include the following functions LAST RUN View the last protocol that was run page 80 SELFTEST Runa self test on the thermal cycler page 81 VERSION View the current instrument firmware version page 82 NAME Enter a name for the thermal cycler page 82 DEFAULTS Change default lid temperature turn off below feature and sample volume page 83 79 1000 Thermal Cycler Manual Advanced Tools and Functions 80 GRADCALC
13. View a temperature gradient based on user defined parameters page 84 CONTRAST Change the instrument s LCD contrast page 86 PORT Change the port used to control the S1000 thermal cycler remotely page 87 View the LAST RUN Tool When you want to see a synopsis of parameters in the last protocol that was run on the S1000 thermal cycler select TOOLS gt LAST RUN NOTE For Dual 48 reaction modules the last run tool toggles to show the last protocol that was run on each block Follow these instructions to view the last protocol that was run 1 Select LAST RUN The LAST RUN option is the default selection when you open the TOOLS option If this option is not selected press the arrow keys to select it Then press ENTER to confirm the selection and continue to the next screen Select LAST RUN LAST FUH SELFTEST WER IOH HAME FORT 2 View the LAST RUN screen The LAST RUN screen shows a synopsis of the parameters in the last protocol that was run Press ENTER to return to the TOOLS option Notice that the parameters listed identify the protocol and any settings that could influence the data from that run The LAST RUN screen displays the following parameters e Name of the protocol e Lid temperature page 33 during the run e Turn off below temperature page 33 for the lid during the run e Sample volume which determines the temperature control mode during page 33 e Total run time e Number of errors detecte
14. e Adding an increment INC an extend EXT a ramp rate change RATE ora signal BEEP parameter to protocol steps by selecting OPTION page 28 e Overview of Temperature Control modes including Calculated and Block modes page 33 e Overview of lid temperature and manual lid settings page 33 e Changing a protocol file using the EDIT option page 34 Creating a Protocol File with the NEw Option Select the NEW option in the main menu to create a new protocol file A protocol file contains protocol steps and instrument instructions to create a cycle of steps NOTE A protocol file can contain up to 99 protocol steps The first step must be a temperature TEMP or gradient GRAD step and the last step must be an END step Before you begin creating a new protocol list the PCR protocol steps and conditions for your PCR experiment including the temperatures and times for each temperature step Then choose the protocol steps page 63 and instrument settings page 64 needed for the protocol For information about optimizing PCR protocols see page 53 NOTE If you are transferring a PCR protocol from another thermal cycler to the S1000 thermal cycler review the suggestions in page 57 This section includes step by step instructions for creating a new PCR protocol on the 1000 thermal cycler These instructions assume you know how to operate the main menu and control panel page 13 19 1000 Thermal Cycler Manual Create and E
15. press the ENTER key to confirm the selection In this example the STD2 file is selected Select a San protocol file Kipi Mies TIP If the folder that contains the file is secure then you must enter the password to rename the file 3 Enter a new protocol file name Press the up and down arrows to select letters and press the number keys to type numbers Then press ENTER to accept the new name and return to the main menu NOTE A protocol file name can contain from one to eight characters The characters are numbers or capital letters Each protocol name must be unique to all folders on the S1000 thermal cycler In this example the new protocol name is STD222 Rename lt MAIH gt Enter a new file name Hew mame 70 Protecting Files in a Folder With the SECURE Function Protecting Files in a Folder With the SECURE Function To protect all the files in a folder select FILES gt SECURE to create a new password or change an existing password Securing a folder protects protocol files stored in that folder To edit move or delete files stored in a secure folder a password must be entered However viewing copying and running protocol files that are located in a secure folder does not require a password To secure a folder follow these instructions 1 Select SECURE To select SECURE press the arrow keys Then press ENTER to confirm the selection and continue to the next screen FOLDERS PROTOCOLS H
16. the lid prevents condensation from forming inside the tubes and plates When running the 1000 s heated lid maintains the lid temperature specified for the protocol being run Without a heated lid water can be lost from the reagents to condensation concentrating the reactants in the tube or plate The thermal cycler s default lid temperature is 105 C NOTE When the block is running an infinite hold at a temperature below the Turn Off Below parameter the lid heater maintains 31 0 C The default Turn Off Below setting is 30 0 C To change the default Turn Off Below parameter select TOOLS gt DEFAULTS 33 1000 Thermal Cycler Manual Create and Edit Protocols Changing an Existing Protocol with the EDIT Option Select the EDIT option when you want to change an existing protocol Here are some reasons to choose the EDIT option e Make small changes to a protocol e Edit a copy of a protocol e Change a copy of a preinstalled protocol Follow these instructions to change an existing protocol 1 Select EDIT from the main menu Press the arrow keys to select EDIT in the main menu then press ENTER to confirm the selection Choose EDIT RUH LIT EH to change HEH FILE a protocol EDIT TOOL Blacks are idle 2 Select a protocol to edit from the file tree Press the arrow keys to select a folder and press the right arrow key select a file within that folder Then press ENTER to continue to the next screen In this ex
17. 4 HH parameters E at a 26 edited mie J ak A c H 30 Step 4 ee a 38 selected OTi 2 36 TIHES 5 ENTER to edit 10 Add an additional parameter to an existing protocol step optional Select a protocol step then choose OPTION to change the parameters of the selected protocol step Then press ENTER to continue to the next screen Lid L16 L lol 26 eL J FOR 4 88 Step 3 z FOUR HH SB selected E FOR 8 SA Choose r Pe Jk OPTION an iT Sari EDIT Choose INC to add an increment to the selected step Then Then press ENTER to continue to the next screen In this example step 3 is selected FOR FOR FOR FOR OTO Choose INC EXT RATE BEEP Enter the increment parameter by pressing the appropriate keys The press ENTER to continue to the next screen r ot D Aan in m T T T oe LTE LTE LT n a ae a ara aoe oe aoe o h af f an 2 a a hay eat Tr i a a a l a z J 39 1000 Thermal Cycler Manual Create and Edit Protocols In this example an increment of 0 sibs is entered FOR FOR FOR F OR i Pd cn an an HNN a ome me LTE LT ara a Cwi ana LTE LTE LTE LT T ae a ay an a amp s ae oe a o E a a bH r ok m a yon Enter paramete n Confirm the parameter Press the right and left arrow keys to select Yes or NO Select Yes to save the changes or select No to return to the beginning of this step Then press ENTER to conti
18. Gene Expression Gateway bio rad com genomics To find local technical support contact your nearest Bio Rad Laboratories office For technical support in the United States and Canada call 1 800 424 6723 toll free phone and select the technical support option 1000 Thermal Cycler Manual ntroduction Writing Conventions Used in This Manual This manual uses the writing conventions shown in Table 6 to quickly provide relevant information Table 6 Writing conventions Convention TIP NOTE WARNING screen message NAME of control panel key Select x Select x gt Y Press X Meaning Provides helpful instructions Provides important information Explains crucial information about a topic that may lead to injury instrument damage data loss Indicates the one or more words on the screen the user should select Indicates a key on the thermal cycler control panel For example these keys have the following names e The ENTER key e The RIGHT ARROW key Select x by using the arrow keys For example select New means Use the arrow keys or to highlight the New option on the screen From menu x select Y For example Select Main gt RUN means Select the RUN option in the main menu Press Xx key on the control panel For example press ENTER means Press the ENTER key on the control panel 1000 Thermal Cycler Manual 2 Get Started The S 1000 thermal cycler
19. Select the file that you want to move Press the arrow keys to select the folder that contains the protocol file Then press the right arrow key to begin selecting the file within that folder Finally press ENTER to confirm the file selection In this example the STD2G protocol is selected Toug lt EWA gt lt GRAHT gt Select file NOTE If the folder is secure then you must enter a password to move the file 3 Select the folder where you want to move the file Press the up and down arrow keys to select the folder where you want to move the file Then press ENTER to confirm the move In this example the file is moved to the EVA folder lt MALH gt Select a folder lt GRAHT gt NOTE If the folder is secure then you must enter a password to move the file Deleting a File With the DELETE Function Select FILES gt DELETE to delete a protocol file when you want to empty a folder or remove extra copies of files To delete a protocol file follow these instructions 68 Copying Moving Deleting and Renaming Protocol 1 Select DELETE To select DELETE press the arrow keys Then press ENTER to confirm the selection and continue to the next screen FOLDERS PROTOCOLS HEW CORT SECURE MVE Select DELETE PELETE JOELETE REHAMNE RENAME 2 Select the file Press the up and down arrow keys to select a folder and then press the right arrow key to select the files in that folder Finally pr
20. a run e Clean the control panel Remove debris on the control panel with a soft cloth and mild soap solution Cleaning this panel prevents damage that will obscure the display NOTE Use of abrasive detergents or rough material will scratch the control panel e Clean the reaction module bay Clean with a damp soft cloth to remove debris and spilled liquids Cleaning this bay allows precise heating and cooling of the reaction block WARNING Never use cleaning solutions that are corrosive to aluminium Avoid scratching the surface of the bay Scratches and damage to this surface interfere with precise thermal control WARNING Never pour water or other solutions in the reaction module bay Wet components can cause electrical shock when the thermal cycler is plugged in e Clean the outside base cover Use a damp cloth or tissue to clean spills off the outside case If needed use a mild soap solution Cleaning the cover prevents corrosion Cleaning the Reaction Modules Clean the S1000 thermal cycler reaction modules on a regular schedule to prevent reagents from accumulating and interfering with the ability of the block to change temperature quickly To clean the reaction module follow these instructions paying careful attention to the warnings WARNING Prevent electrical shock Always remove the reaction module from the thermal cycler base before cleaning it e Clean the cooling fins Remove dust with a soft brush or damp cl
21. a folder Once the folder is selected the right side of the screen lists all the protocol files in that folder 75 1000 Thermal Cycler Manual Protocol Files and Folders 76 In this example the MAIN folder is selected PROTOCOLS 3 Select a protocol file to view Press the right arrow keys to begin selecting a protocol file within the folder Then press the up and down arrows to select a protocol file Finally press ENTER to continue to the next screen NOTE If the folder lists more than 6 files view the full list of files by pressing the down arrow key In this example the ITAQOFST file is selected PROTOCOLS LET lt EWA gt lt GRAHT gt al Fes Select protocol file Ts mfn E E iz RE RF A T IF IF IT 4 View the protocol file Press the ENTER key or the up and down arrow keys to scroll through the protocol file until the END step In this example all six steps in the protocol fit on one screen Lii gw File name ITAGQFET file 1 4s H r AS gs 15 Vhs Steps in 5 j 3 l rotocol z ad e i z 8 E oe i E T E View the end of the protocol q E r Final step in file Or ob al Po Viewing Protocol Files With the VIEW Option 5 Press ENTER or CANCEL again to finish viewing the file Press the ENTER key to finish viewing the protocol and return to the main menu 77 1000 Thermal Cycler Manual Protocol Files and Folders 78
22. can reduce the total run time by a third In contrast running the same protocol on a thermal cycler with a faster ramp rate only cuts minutes from the total run time Furthermore optimizing the protocol for speed can also result in better PCR results TIP Automatically create a protocol that is optimized for speed by using the Protocol Autowriter on the C1000 thermal cycler or in 1000 series software Run the S1000 thermal cycler under the control of a C1000 thermal cycler and use the Protocol Autowriter to write protocols to run on both thermal cyclers page 88 53 1000 Thermal Cycler Manual Optimizing PCR With the S1000 Thermal Cycler 54 To optimize a PCR protocol to run faster by using the features of the S1000 thermal cycler follow these guidelines Shorten the denaturing step The initial denaturation step page 61 requires a longer hold time than other denaturation steps during the cycle This difference is due to the activation of the polymerase and the longer initial DNA template Once the PCR target is amplified the amplicons then serve as shorter templates that are easier to denature during cycling To shorten the denaturation step enter a hold time of 1 second for PCR products that are less than 500 base pairs bp Then test this shorter hold time to verify that 1 second denaturation is sufficient to produce amplicons Alternatively add an increment page 28 to the denaturation step to test for the best hold time
23. example of a file tree In this example the MAIN folder is selected and the protocol files in that folder are listed on the right side of the screen This protocol list includes the preinstalled protocol files and user created protocol files and the selected protocol file is ITAQFST In this example the folders named EVA The File Tree and GRANT are user created folders Notice that all folder names display with angle brackets lt and gt surrounding the name User created Selected folder protocol file User folders GRAHT gt Preinstalled protocols Selected file ee do oi en TT a i I T NOTE If a folder contains more than six protocols then it is necessary to scroll down to see all the protocols To scroll through a folder press the arrow keys The MAIN folder always contains the preinstalled protocols page 101 The preinstalled files in the main menu can be run or copied by any user When the main menu becomes full of files it is difficult to find a specific protocol Move any user created protocol files to another folder for easy access page 65 17 1000 Thermal Cycler Manual Get Started 18 1000 Thermal Cycler Manual 3 Create and Edit Protocols The main menu provides quick access to the options needed to create and edit protocol files This chapter provides instructions about using the NEW EDIT and FILES options e Creating a new protocol file by selecting the NEW option below
24. hold time and cycle In this example the STD2 protocol is running Step 2 and the current cycle 46 Monitoring a Run with the SCREEN Key is 2 The current temperature is 95 0 C and the hold time is currently 30 seconds Current step and parameters Current temperature Current hold time 2 The graphical screen This screen displays a graph that approximates the relationship of the target temperatures in each step in the protocol Each step is listed by temperature only To display the current step a highlighted temperature and flashing bar display the temperature in the graph The current hold time blinks above the graph while counting up to the total hold time In this example the protocol has five steps including one GOTO step Step 2 is currently running and the temperature is 95 0 C Protocol step Hold time for current step counts up by temperature Target temperature J 5 Fi ites Flashing thick for the current step zis bar shows current step 3B x To display the GOTO step a dotted line in the graph links the steps within the GOTO repeat The total number of repeats in the GOTO step is displayed below the dotted line In this example the GOTO repeat begins with step 2 and repeats 30x The GOTO includes steps 2 and 3 q5 8 8 29 Step 2 Steps that repeat eS in the GOTO GOTO Step 3 repeat Number of repeat In GOTO step Dotted line shows protocol steps that repeat
25. in the COTO step 47 1000 Thermal Cycler Manual Run Existing Protocols 3 The Time Remaining screen This screen displays the amount of time remaining until the end of the protocol In this example the protocol has 33 minutes and 24 seconds remaining in the run Amount of time remaining for protocol run hr min sec Time Remaining NOTE The calculation for the time remaining includes the hold time for each protocol step and the time it takes to ramp to each target temperature Additionally infinite holds are ignored in the calculation NOTE Dual blocks have the time remaining and other information for each block Pausing and Resuming a Run with the PAUSE Key When you want to temporarily stop a protocol use the PAUSE key During a pause the thermal cycler maintains the block temperature at the current target temperature If the block temperature has not reached the target temperature then the thermal cycler will continue to heat or cool until it reaches that target temperature WARNING Pausing a protocol can alter the results of your PCR experiment When you pause a protocol during a temperature step you create a longer hold time for that step To pause and resume a protocol follow these instructions 1 Press PAUSE to stop the protocol Press the PAUSE key Notice that the time display shows PAUSE to indicate that the protocol is paused In this example step 1 in cycle 1 is paused a
26. is easy to set up and run insert the reaction module plug in the thermal cycler base then begin a PCR experiment This chapter describes how to set up and operate the S1000 thermal cycler Refer to the following sections for detailed instructions e Setting up the S1000 thermal cycler below e Introducing the 1000 thermal cycler base and interchangeable reaction modules page 7 e Choosing and inserting a reaction module page 9 e Operating the lid of the reaction module page 12 e Operating the control panel page 13 e Selecting files and folders from the file tree page 16 Setting Up the 1000 Thermal Cycler This section provides an overview of the steps needed to safely unpack and set up the 1000 thermal cycler Follow these instructions to prepare to run the thermal cycler 1 Unpack the S1000 thermal cycler base and reaction module The S1000 thermal cycler base package includes the thermal cycler base a power cord quick guide and this manual Keep all packaging to safely move this instrument Place the thermal cycler base in the desired location NOTE A thermal cycler requires sufficient cool air flow to run precisely To check for sufficient air flow before running the instrument see page 91 2 Insert a reaction module The reaction module fits into the thermal cycler bay and locks in place for operation For detailed instructions about loading the reaction module see page 10 3 Plug in the power cord an
27. of a protocol a protocol can only contain one END step To enter an END step press the arrow keys to select END Then press ENTER to continue to the next screen In this example the END step type is selected Choose END step TI oe mar ey nh GRAD ENTER wt ge on wi pS pe A Confirm the END step Press the arrow keys to select Yes or No Select Yes to accept the protocol step parameters or select No to return to the beginning and select a different protocol step Confirm END step 11 Save the new protocol file in an existing folder in the file tree If there are no user created folders i e there is only one folder MAIN the file tree folder screen will not appear Press the arrow keys to choose a folder from the list of folders on the S1000 thermal cycler Store the protocol file in a folder that you created or in the MAIN folder Press ENTER to save the protocol file in the selected folder and return to the main menu In this example the selected folder is called GRANT Save SIDZGRAD lt MALH Selected HAJI E folder GRAHT 3 TIP If you store the protocol in the MAIN folder you can move it to a specific folder with the FILES gt MOVE page 67 To run a protocol follow the instructions in Running a Protocol With the RUN Option on page 41 27 1000 Thermal Cycler Manual Create and Edit Protocols Adding INC EXT RATE and BEEP Parameters 28 A new temperatu
28. press the CANCEL key In this example the incubation temperature is 95 C Incubation temperature Incubate af 51 1000 Thermal Cycler Manual Run Existing Protocols 52 4 Start the incubation Press the ENTER key to start the incubation In this example the block is incubating at 95 0 C RUH WT EH HEH ELEES BRET TOOLS Incubation temperature Inczukbating at eS TIP During incubation use any options in the main menu except RUN You cannot start a run on a block that is running an incubation 5 Press CANCEL to stop the incubation Press the CANCEL key to stop the incubation then select YES or No Select YES to stop the incubation or select No to continue the incubation Finally press ENTER to confirm the selection and return to the main menu In this example YES is selected WT EH PLCE TOOL Select YES to cance the incubation Cancel run Ho 6 View the incubation parameters To see the parameters of the incubation press the SCREEN key three times to view the LAST RUN screen To view the parameters of the run press the SCREEN key twice to view the LAST RUN screen In this example the lid temperature was 100 C the incubation ran for 8 seconds and the version of the firmware at the time of the incubation was 0 2 0 96 LAST RUH INCUBATE ELD 146 30 VOLUME 18 TOTRBL TIME 0 H9 ERRORS Horg MEREION 85 2 0 966 1000 Thermal Cycler Manual 5 Optimizing PCR With the 1000 Th
29. protocol file to another folder page 67 65 1000 Thermal Cycler Manual Protocol Files and Folders e Select DELETE to delete a protocol file page 68 e Select RENAME to rename a protocol file page 69 Copying a File With the Copy Function To begin copying a protocol file select FILES gt COPY The COPY function makes a copy of the existing file and requires you to enter a new file name for the copy You can also move the protocol to a new folder For example move secured protocols or preinstalled protocols to your folder by copying them To copy a protocol file follow these instructions 1 Select COPY To select COPY press the arrow keys then press ENTER to confirm the selection and continue to the next screen FOLDERS PROTOCOLS sian i SECURE Select COPY CELETE DELETE REHAME REHAME 2 Select the protocol file to copy Press the arrow keys to select the folder that contains the protocol file then press the right arrow key to select a file Press ENTER to continue to the next screen In this example the STD2 file is selected in the MAIN folder LOH ts lt EWA gt LOHG Select a protocol sams RAHT gt TO2 file to copy IPRFSEB IPRFAiKB ITAGFST 3 Select the folder where you want to store the copy Press the arrow keys to select a folder where you want to store the copied protocol file Press ENTER to confirm the selection 66 Copying Moving Deleting and Renaming Protocol In
30. reaction module The following sections describe important parts of the thermal cycler and reaction modules e Front view of the base without a reaction module below e Top view of the base with reaction module page 7 e Back view base with reaction module page 8 Front View without Reaction Module This front view of the thermal cycler base shows a control panel and reaction module The following image shows major features of the thermal cycler base including the control panel LCD screen and air vents Top View with Reaction Module This top view shows the reaction module docked in the bay of the thermal cycler base The following shows major features of the reaction module and thermal cycler base For more information about reaction modules see page 9 Reaction module Module locking bar 1000 Thermal Cycler Manual Get Started Back View with Reaction Module The following shows the major components on the back of the thermal cycler with a reaction module including the power switch data ports and reaction module Back of reaction Locking bar module Power switch __ Power input Back panel of base Data ports Cooling vents The back panel includes data ports that transfer data to and from the thermal cycler e USB B port for connecting the S1000 thermal cycler to a C1000 thermal cycler e USB A ports for connecting to another S1000 thermal cycler that is connected to a C1000 thermal cycler or to a US
31. sec or Repeats 95 60 increment at 0 5 C iai Infinite hold OO ifaq Polymerase Protocols iTfaq DNA polymerase is an antibody mediated hot start polymerase that is suitable for both PCR and real time The iTaq protocols run PCR using the optimal parameters 102 iProof Polymerase Protocols for this polymerase and associated buffers ifaq polymerase is designed to be activated during the first step at 98 C and to amplify small to medium size templates Table 17 iTaq Protocol iITAQ FST To Run Fast PCR Target Temperature iProof Polymerase Protocols iProof DNA polymerase is a high fidelity polymerase that is designed to quickly and precisely amplify long targets using a proofreading enzyme combined with a DNA binding protein These iProof protocols include optimal parameters for this enzyme and associated buffers including an initial 95 C step to activate the enzyme and a final long extension step Each of these protocols is adjusted for a distinct range of target sizes Table 18 lists the parameters in each protocol Table 18 iProof Protocol IPRF1KB IPRF8KB To amplify 1 kb or To amplify 8 kb or smaller targets fast smaller targets Infinite Infinite hold hold OO OO 72 IPRF15KB To amplify 15 kb or smaller targets Infinite hold CO 103 1000 Thermal Cycler Manual Appendix A Preinstalled Protocols Table 18 iProof Protocol IScript Polymerase Protocol Table 19 IScri
32. target temperature add the BEEP parameter to the step page 28 When the sample reaches the target temperature the S1000 thermal cycler begins to count up to the total hold time for that step TIP Once the run starts you can view the progress of the run in several screens To view these screens press the SCREEN key Each time you press the SCREEN key the thermal cycler displays the next screen in a series that displays the status of the run page 46 After running the first cycle the screen shows the GoTo repeats In this example the S1000 thermal cycler is running Cycle 2 and the total number of cycles is S15 Total number Current cycle of cycles NOTE The number of GOTO repeats does not include the first cycle the total number of cycles is the number of repeats plus one When the protocol runs the final temperature step it completes the protocol In this example the final temperature step is Step 4 Final temperature step Final cycle returns to first cycle 45 1000 Thermal Cycler Manual Run Existing Protocols 8 View the Protocol complete screen When the protocol is finished the S1000 thermal cycler displays the Protocol complete screen After viewing this screen press ENTER to return to the main menu In this example the ITAQOFST protocol ran for 10 minutes and 53 seconds PROTOCOL COMPLETE Total run time Total Time 18 53 9 View the LAST RUN screen optional Once the 1000 ther
33. the instructions in Step 5 page 22 to continue entering temperature steps In this example four steps were entered Entered step Choose new step type NOTE A protocol can contain up to 99 protocol steps The first step must be a temperature TEMP step and the last step must be an END step 8 Enter a GOTO step immediately after the set of steps that cycle optional The GoTo step creates the cycle in the PCR protocol For more information about how the GoTo step creates a cycle see Protocol Steps page 61 Press the arrow keys to choose GOTO Press ENTER to continue to the next screen In this example step 5 will be a GOTO step qzw ce Coe ad ee aa 5 d 72 p FOR 0 Choose GOTO 4F GRAD T oa G NOTE The GoTo step cannot be the first or the last step in the protocol A protocol always starts with a TEMP or GRAD step and ends with an END step Enter the step number for the first step in the COTO repeats Press the number keys to enter a step number Then press ENTER to continue to the next screen 25 1000 Thermal Cycler Manual Create and Edit Protocols 26 In this example the first step is 2 The GOTO step instructs the thermal cycler to return to step 2 and repeat all the steps between step 2 and step 5 the GOTO step Steps that repeat during GOTO step E First step i GOTO repeats Enter the number between 1 and 9999 for the additional repeats ADDTN
34. well block is 71 4 C FROHT ROH Temperature Rows in 96 well in row D block A H sis aia afa Gen T A e a h i h A mipmap ty J B iN iN J D pe LB L H Ei C i E F G H BACE ROL NOTE The temperature in the middle rows is estimated The estimate is based on the temperatures in the front and back rows and on the number of rows in the block Changing Screen View with the CONTRAST Tool Select the TOOLS gt CONTRAST tool to change the contrast of the LCD when you want to improve the visibility of characters displayed on the screens To change the CONTRAST follow these instructions Select CONTRAST To select CONTRAST press the arrow keys Then press ENTER to continue to the next screen TUULA LH T RUH DEFAULT SELFIE SI GbRAULALL Select CONTRAST Mit aes any CONTRAST HAME PUR T Adjust the contrast Press the right and left arrow keys to adjust the contrast Current contrast O M EHTER TO ACCEPT CAHCEL Press the left arrow key to decrease the contrast The TOOLS Options In this example the contrast is decreased Lower contras Decrease contras EHTER TO ACCEPT CAHCEL TO REJECT Press the right arrow key to increase the contrast In this example the contrast is increased Increase contrast Increase contras EHTER TO ACCEPT CAHCEL TO RETECT 3 Accept or reject the contrast Press the ENTER key to accept the changes and
35. with heated pressure plate ET Thermal block_ j 96 well Fast r i Reaction module lid zx LED 1000 Thermal Cycler Manual Get Started Inserting and Removing a Reaction Module Inserting and removing a reaction module is quick and easy When inserted a reaction module locks into the thermal cycler base for safe operation This section includes instructions for inserting and removing a reaction module INSERTING A REACTION MODULE To insert a reaction module follow these instructions 1 Align the reaction module in the bay Lower the locking bar to the unlocked position Then lift the module into the reaction module bay and leave about 2 cm of space in front Locking bar unlocked down 2 Lock the reaction module in place Pull up the locking bar to push the module forward locking it in place Locking bar locked up position TIP Store the reaction module in the base when not in use 10 Interchangeable Reaction Modules REMOVING A REACTION MODULE Follow these instructions to remove a reaction module from the base 1 Unlock the reaction module Push down the locking bar to unlock the module Locking bar down unlocked 2 Lift the reaction module out of the bay Grasp the front and back of reaction module and lift it up and out of the bay 3 Store the reaction module on a clean flat surface If you choose not to store the reaction module in the base be s
36. 1000 offers fast thermodynamics for shorter run times and expedited results e Reliable Results with Precise Thermal Control Sample block uniformity of 0 4 C at 90 C after 10 seconds with an accuracy of 0 2 C adjustable lid temperature to further control reaction temperature and prevent evaporation e Thermal Gradients Optimize reaction performance using temperature gradients across sample blocks e Flexible Vessel Format with Interchangeable Reaction Modules Simply change reaction modules when you need 48 96 or 384 well blocks with the same thermal cycler base Uniform and repeatable microplate sealing with adjustable lid force e Pre installed Customizable Protocols for Use as Templates Speed protocol writing by customizing pre loaded protocols e Ready for High Throughput Use remote commands to control up to 32 thermal cyclers for automated robotic systems Bio Rad Resources and References Bio Rad Resources and References Bio Rad Laboratories provides many resources for scientists The following websites contain useful information about running PCR experiments e Gene Expression Gateway bio rad com genomics This site provides rich technical resources on a wide variety of methods and applications related to PCR and real time PCR This site also features tools citations technical support and troubleshooting resources e Life Science Research web site discover bio rad com This site includes links to tech
37. 33 In this example the default sample volume for the thermal cycler is 10 uL and the mode is Calculated The thermal cycler will use the sample temperature to determine when it reaches the target temperature for a step Run ITAQFST on A Enter sample volume Sample Vol i ony RUN UI EW TIP When running a dual block the message is Run ITAQFST on A where ITAQFST is the protocol and A is the block Press the numeric keys to enter a new sample volume then press ENTER to continue to the next screen In this example the sample volume is changed to 25 uL Run ITAQFST on A Change E sample volume sample Nol E SAL RUN VI EW 5 Select VIEW to review the protocol before beginning the run optional Press the right arrow key to select VIEW then press ENTER to confirm the selection and review the protocol Rum ITAQFST 7 sample Vol 2517 Select VIEW 43 1000 Thermal Cycler Manual Run Existing Protocols 44 While reviewing the protocol press ENTER to scroll down through the steps in the protocol When you reach the last step in the protocol press ENTER again to exit In this example the screen shows the steps in the ITAQOFST protocol Lid 188 C View the ITAOFST H Wal 28pL protocol r a D D bm p o J SS aa M M x r L Protocol steps a ee ra ey me ee a i pe a a H pa Rot T J 5 T J Select RUN to begin the run Press the arrow keys to s
38. B B Port 88 Connecting Indirectly to a C1000 Thermal Cycler via the 1000 USB A Port 89 Operating the 1000 Thermal Cycler While Under C1000 Thermal Cycler Control 90 Automated Control for Robotic Systems 00000 eee ees 90 Chapter 8 Maintenance amp Troubleshooting 02000000eeae 91 Cleaning and Maintaining the S1000 Thermal Cycler 91 Cleaning the 1000 thermal cycler Base 91 Cleaning the Reaction Modules 92 Maintaining Sufficient Air Flow 0 000 e ee eee ees 93 Testing for Sufficient Airflow 93 Fixing Insufficient Airflow 94 Replacing Fuses 94 Troubleshooting ica eee ech beanies eee Be Greed 95 Restarting After a Power Failure 95 Warning and Error Messages 95 scenes erase tere ete Sanat Oued ie ete arr eee aod Silenced are wee anaes Gea eee eee ee 99 Appendix A Preinstalled ProtocolS 0 0000 eee eee 101 Standard PrOloCOlS sisi kde aries he ee eee ee eee ae 101 TOUGNGOWN PIOlOCOls i os c0 cc0es 662i bee eee eeee when eae ewes 102 iTaq Polymerase Protocols 0 0c e eee ees 102 iProof Polymerase ProtocolS 2 000 cece nents 103 IScript Polymerase Protocol 00 ccc eee eee ees 104 Nested Primer PrOlOCOlS 63 46 oii ed BS 4S Sek oe Se et 104 Appendix B Operating Specifications 0 00000 eee eee eee 107 Instrument Operating Specifications 0000 cee eee eee 107 Gradient Specifications 0000 c eee ee 108 Appendix C Catalog Numbers
39. B hub page 88 e Test port used for service testing only USB B port USB A ports Power switch Powe input Fuses Cooling vents Test port Interchangeable Reaction Modules Interchangeable Reaction Modules The S1000 thermal cycler can run any of the 1000 series reaction modules The reaction modules include fully adjustable heated lids that are capable of reliably running with a broad range of plastics Table 7 1000 series reaction module size and limits Recommended sample Number of wells Number of blocks ae volume upper limit Dual 48 10 50 uL 50 uL limit 96 10 50 uL 50 uL limit 384 3 30 uL 30 uL limit CFX 96 Real Time CFX 384 Real Time 10 50 uL 50 uL limit 3 30 uL 30 uL limit All reaction modules are compatible with a large variety of PCR reaction vessels in addition to the tested microplates tubes caps and sealing tape listed on page 58 All reaction modules include the features shown in the following image e Heated inner lid Set the lid temperature for each protocol The lid also applies the force that seals reaction vessels e Lid force knob Adjust the knob to obtain appropriate lid force to seal the reaction vessel e LED Turns on to show which block is selected and or in use e Locking lever Secures the reaction module to the base during operation e Sample Thermal block Holds sample vessels and controls the temperature throughout the run Lid locking lever Inner lid
40. C The target temperature for step 3 will increase by 0 5 C each cycle Enter increment parameter Confirm the step parameters Press the arrow keys to select YES or No Select YES and press ENTER to confirm the parameters for protocol step 3 Select No and press ENTER to change the parameters 29 1000 Thermal Cycler Manual Create and Edit Protocols Extending EXT the Hold Time in a Temperature Step The EXT parameter changes the hold time for a temperature or gradient temperature step The extension increases or decreases the hold time with every cycle To add an extension follow these instructions 1 Choose OPTION after entering the initial temperature step parameters Press the right and left arrow keys to choose OPTION Then press ENTER to continue to the next screen In this example the OPTION will be added to step 3 parameters Step 3 Choose OPTION OPTIOH NOTE The EXT option must be added to a step within a GOTO repeat in order to extend with each cycle of the reaction 2 Choose EXT to extend the hold time each cycle Press the right and left arrow keys to select the EXT parameter Then press ENTER to continue to the next screen ee LT ea a n meet oe Choose EXT wel RATE BEEP 3 Enter the extension time in seconds Press the number keys to type a parameter To decrease the hold time each cycle press the CANCEL key to enter a minus sign Then press the ENTER key to contin
41. ED PLEASE CALL BIO RAD FOR SERVICE BLOCK SENSOR 4 FAILED PLEASE CALL BIO RAD FOR SERVICE BLOCK SENSOR 5 FAILED PLEASE CALL BIO RAD FOR SERVICE LEFT LID SENSOR FAILED PLEASE CALL BIO RAD FOR SERVICE RIGHT LID SENSOR FAILED PLEASE CALL BIO RAD FOR SERVICE LEFT HEATSINK SENSOR FAILED PLEASE CALL BIO RAD FOR SERVICE RIGHT HEATSINK SENSOR FAILED Cause Block failed to achieve target in the estimated time Lid failed to achieve target in the estimated time Block failed to achieve gradient in the estimated time singles only If the right and left lid heater channels deviate from each other by more than 5 C the lid is shut off Block sensor 0 has failed and the protocol was terminated Block sensor 1 has failed and the protocol was terminated Block sensor 2 has failed and the protocol was terminated Block sensor 3 has failed and the protocol was terminated Block sensor 4 has failed and the protocol was terminated Block sensor 5 has failed and the protocol was terminated Left lid sensor has failed If a dual protocol terminated If a single and BOTH lid sensors failed protocol terminated and block sent to 4 C Right lid sensor has failed If a dual protocol terminated If a single and BOTH lid sensors failed protocol terminated and block sent to 4 C Left heatsink sensor has failed system using average of amplifier temperatures to continue Right heatsink sensor h
42. EH COPY Select SECURE Mowe DELETE DELETE RENAME FENAME 2 Select a folder to secure Press the up and down arrow keys to select the folder you want to secure with a password In this example the GRANT folder is selected SCcure TILE lt EV gt Select a folder GRAHT gt Henu ee de ee TIP When you want to change a password follow the same instructions you used to create the password When you change a password you will be required to enter the original password before you can change it NOTE If the password is lost then protocols in the secured folder cannot be deleted moved or edited Furthermore the folder cannot be deleted However the protocols in a secure folder can be copied to another folder 3 Enter a new password Press the number keys to type a number Then press ENTER to confirm the password and return to the main menu 71 1000 Thermal Cycler Manual Protocol Files and Folders In this example the password Is 222 lt MAIH gt EWA gt lt GRAHT gt Enter a new password os SLOT fa 222 NOTE A password can contain from one to four numbers and cannot contain letters TIP To disable security for a folder repeat steps 1 3 and specify a blank new password Creating Deleting and Renaming Folders 72 The S1000 thermal cycler stores protocol files in the MAIN folder by default If you want to store the files in another location a folder must be created This section
43. FCC WARNINGS AND NOTES Warning Changes or modifications to this unit not expressly approved by the party responsible for compliance could void the user s authority to operate the equipment Note This equipment has been tested and found to comply with the limits for a Class A digital device pursuant to part 15 of the FCC Rules These limits are designed to provide reasonable protection against harmful interference when the equipment is operated in a commercial environment This equipment generates uses and can radiate radio frequency energy and if not installed and used in accordance with the instruction manual may cause harmful interference to radio communications Operation of this equipment in a residential area is likely to cause harmful interference in which case the user will be required to correct the interference at his own expense Note regarding FCC compliance Although this design of instrument has been tested and found to comply with Part 15 Subpart B of the FCC Rules for a Class A digital device please note that this compliance is voluntary for the instrument qualifies as an exempted device under 47 CFR 15 103 c in regard to the cited FCC regulations in effect at the time of manufacture Note regarding Canadian EMC compliance Le present appareil numerique n emet pas de bruits radioelectrique depassant les limites applicables aux appareils numeriques de class A prescrites dans le reglement sur le brouillage radi
44. I u a T A NTER to edit 6 Choose an editing method This screen displays four editing methods Press the arrow keys from this screen to choose one of these methods e Choose INS to insert a step above the selected protocol step see Step 7 on page 36 e Choose DEL to delete the selected protocol step see Step 8 on page 37 e Choose EDIT to change the parameters in the selected protocol step see Step 9 on page 38 e Choose OPTION to add more parameters to the selected protocol step see Step 10 on page 39 TIP To replace a temperature step with a gradient step choose INS to insert the gradient step and then choose DEL to delete the temperature step To stop editing the protocol press the CANCEL key several times If asked Save Changes press YES to save Press No to discard edits 35 1000 Thermal Cycler Manual Create and Edit Protocols In this example INS is selected to insert a step above the selected step L a 118 G Wol z p oR 2 O akale Ei J E oto L j a Jin h iJ hd i i a T man E m mM A _ E oie Choose a methoc 7 Insert a protocol step optional Choose INS to insert a protocol step above the selected step Press ENTER to continue to the next screen In this example a new step will be inserted above step 4 Lid 119 C ial i 45 8 FOR f 45 8 FoR 3 29 87 FUR Selected step 4 G TE EA 5 10 0 F R Choose INS IH
45. ITAQFST Next step ee a a E TE p Temp T aj s Cycl 49 1000 Thermal Cycler Manual Run Existing Protocols Stopping a Run With the CANCEL Key To stop a protocol press the CANCEL key You can stop a protocol at any point during the run When a protocol is cancelled the block immediately stops changing temperature To stop a protocol follow these instructions 1 Press CANCEL and select YES Press the CANCEL key then press the right and left arrow key to select YES or No Finally press the ENTER key to confirm the selection and continue to the next screen In this example YES is selected Running on ITAGFET Step 3 65 87 amp Temp TS 4 Repeat 1 Select YES to cancel MERTI E SUE TIP For the Dual 48 reaction module if you press CANCEL while viewing the main menu then you need to select the block that is running the protocol you want to cancel before proceeding 2 Press ENTER to stop the protocol and view the PROTOCOL CANCELLED screen Press the ENTER key to confirm that you want to stop the protocol The 1000 thermal cycler displays the PROTOCOL CANCELLED screen This screen displays the total time that the protocol ran In this example the protocol ran 2 minutes and 57 seconds PROTOCOL CAHCELLED Total run time Total Time Si 3 Press ENTER again to return to the main menu Incubating Samples With the INCUBATE Key To incubate samples at a single temperature pres
46. In this example the option will be applied to step 7 Choose BEEP IHC EXT RATE Haag NOTE The BEEP parameter can only be added to a temperature step 3 Confirm the step parameters Press the arrow keys to select YES or No Select YES to confirm the selection and finish entering the parameters for step 3 Select No and change the parameters for this protocol step Then press ENTER to continue to the next screen Setting Temperature Control Mode The S1000 thermal cycler uses one of two temperature control modes to determine when the sample reaches the target temperature The thermal cycler determines the sample temperature by utilizing one of the following temperature control modes e Calculated mode When you enter a sample volume between 1 and 50 uL or 30 uL for 384 well reaction modules the thermal cycler calculates the sample temperature based on the sample volume This mode is the recommended temperature control mode because it most accurately represents the actual sample temperature e Block mode When you enter a sample volume of zero 0 uL the thermal cycler assumes the sample temperature is the same as the measured block temperature To select a temperature control mode while creating a new protocol see Step 4 page 21 To select a temperature control mode when starting a protocol see Step 4 page 43 Choosing Lid Temperature The adjustable heated lid allows the user to control lid temperature and force Heating
47. L REPEATS in the GOTO step Then press ENTER to continue to the next screen NOTE The GoTo step adds additional cycles to the PCR protocol The first cycle is not included in the GoTo step For example to run a PCR protocol with 31 cycles enter 30 repeats in the GOTO step In this example the number of repeats is 30 and the total number of cycles is 31 Hu Steps that repeat in the GOTO step Number of GOTO 2 additional repeat OTHL REFEAT in GOTO step Confirm the parameters Press the arrow keys to select Yes or No Select Yes to accept the GOTO step parameters or select No to return to the beginning of this step and change the GoTo step parameters Then press ENTER to continue to the next screen FUR FOR Confirm GOTO parameter l 3 a K 9 Enter the remaining steps in the protocol file Enter the remaining steps by choosing the step type and adding parameters Then press ENTER to continue to the next screen NOTE A protocol can contain up to 98 TEMP GRAD and GOTO steps The first step is always a TEMP or GRAD step The last step is always an END step TIP To instruct the thermal cycler to emit a sound at the end of the protocol include a BEEP option in the final temperature step page 32 Creating a Protocol File with the NEw Option 10 Enter an END step to finish the protocol file The END step instructs the thermal cycler to finish the protocol file NOTE The END step must be the last step
48. Menu Closing the Lid and Setting Lid Force To close the lid follow these instructions 1 Close the lid Insert the reaction tubes or plate into the reaction module Push the lever down and back to close the lid and lock it in place Lever Lid lock locked 2 Adjust lid force Turn the lid force knob clockwise to the right to increase the lid force Turn counter clockwise to the left to decrease lid force TIP First turn the lid force knob clockwise until the lid makes contact with your plate or tubes it will feel slightly tight Then turn the knob 1 2 turn past contact if using a tape seal or 1 4 turn past contact if using strip caps or individual tubes Lid force knob NOTE Do not turn the lid force knob more than a 1 2 turn after the inner plate touches the vessels Excessive force is not necessary Control Panel Keys and Main Menu The control panel on the S1000 thermal cycler provides access to all the functions needed to run the thermal cycler The control panel includes the following e Liquid crystal display LCD Displays the main menu and other screens e Keypad amp Navigation Keys Enter commands with these keys page 15 13 1000 Thermal Cycler Manual Get Started LCD showing Numeric and symbol keys main menu Navigation keys HENDEL SCREEN x cat key a De 1000 Thermal Cycler 14 Control Panel Keys and Main Menu Control Panel Keys Table 8 Functions o
49. OR SERVICE PLEASE CHECK AIRFLOW PS OVERHEATING PLEASE CALL BIO RAD FOR SERVICE Cause Displayed when a machine running a protocol has been turned off either intentionally or due to a power outage and then turned on again Reaction module has exceeded maximum temperature of 107 5 C or sensor has a malfunction and is not measuring temperature accurately Protocol terminated Heatsink temperature has exceeded 75 C Protocol terminated Amplifier 1 temperature has exceeded 85 C Protocol terminated All block sensors have failed see below for failure criteria Protocol terminated Power Supply temperature has exceeded 85 C Protocol terminated Lid sensor has failed during lid preheat Protocol terminated Heatsink has exceeded 70 C System beeps and displays error Amp temp has exceeded 80 C System beeps and displays error Power Supply temperature has exceeded 80 C System beeps and displays error Troubleshooting Table 14 Warning and error message solutions Message PLEASE CALL BIO RAD FOR SERVICE SLOW BLOCK CYCLING PLEASE CALL BIO RAD FOR SERVICE SLOW LID CYCLING PLEASE CALL BIO RAD FOR SERVICE SLOW GRADIENT PLEASE CALL BIO RAD FOR SERVICE HEATED LID FAILED PLEASE CALL BIO RAD FOR SERVICE BLOCK SENSOR 0 FAILED PLEASE CALL BIO RAD FOR SERVICE BLOCK SENSOR 1 FAILED PLEASE CALL BIO RAD FOR SERVICE BLOCK SENSOR 2 FAILED PLEASE CALL BIO RAD FOR SERVICE BLOCK SENSOR 3 FAIL
50. S EDIT OPTIOH Choose the type of protocol step to insert then press ENTER to continue to the next screen In this example a temperature step TEMP is selected Choose step type EHTER to gdit Enter the step parameters Then press ENTER to confirm each parameter In this example the target temperature of 72 C is entered 35 8 Inserted step GOTO Enter 1 At temperature FHO parameter TEMP L Enter more step parameters by choosing Option optional For example add an increment or extension to this temperature step For more instructions about entering additional parameters to a step see Adding INC EXT RATE and BEEP Parameters on page 28 36 Changing an Existing Protocol with the EDIT Option In this example OP TION Is selected to add parameters to step 4 wd m CL D E biun it E Te ee oe os ate r Inserted step E en es m Choose OPTION T z K 2a mf ay Ho OS Enter the parameters of the new step Then press ENTER to confirm each parameter In this example the target temperature of 72 C is entered wI FT CL D E m T aN E e hon Tt E ToD ee E Inserted step fo l I i A DDD CE fs Confirm parameter colle 3 m Hoo ption 8 Delete a protocol step optional Choose DEL to delete the selected protocol step In this example
51. S1000 thermal cycler Plug the other end of the cable into a USB A port on the previous 1000 thermal cycler the S1000 connected directly to the C1000 4 Connect an additional S1000 indirectly to the C1000 By repeating steps 1 3 you may connect up to three 1000s indirectly to the same C1000 5 Open the instrument tree Open the instrument tree on the C1000 thermal cycler or in the 1000 series software 6 Select the S1000 thermal cycler Select the S1000 thermal cycler by serial number or name If the S1000 thermal cycler has a name page 82 then the name will display instead of the serial number 89 1000 Thermal Cycler Manual Advanced Tools and Functions Operating the 1000 Thermal Cycler While Under C1000 Thermal Cycler Control When the S1000 thermal cycler is under the control of the C1000 thermal cycler it is in Semi lock down mode In Semi lock down mode the S1000 thermal cycler will not respond when control panel keys are pressed However the following keys function on the control panel e SCREEN key Press this key to access the Running Screen Graphical Screen and Time Remaining Screen e PAUSE key Press this key to temporarily stop a protocol that is currently running on the S1000 thermal cycler e ENTER key Press this key to skip a step e CANCEL key Press this key to cancel a protocol that is currently running on the 1000 thermal cycler Automated Control for Robotic Systems 90 The
52. S1000 thermal cycler is capable of responding to robotic control commands in a high throughput workflow To connect the 1000 thermal cycler to a robotic system use either a USB or serial RS232 port Choose a port by selecting the PORT tool page 87 and then selecting the port Once connected robotic commands automate basic operations of the S1000 thermal cycler For information about these commands download the Automation Guide from the Gene Expression Gateway bio rad com genomics This guide includes instructions and commands for running S1000 thermal cyclers under the control of a robotic system For additional information about running the thermal cyclers in the Bio Rad 1000 series within an automated environment refer to the C1000 thermal cycler manual 1000 Thermal Cycler Manual 8 Maintenance amp Troubleshooting The following chapter contains important suggestions for maintaining and troubleshooting the S1000 thermal cycler Cleaning and maintaining the 1000 thermal cycler for optimal safety and performance below Maintaining sufficient air flow around the thermal cycler to optimize thermal control while running PCR page 93 Troubleshooting options including how to interpret and respond to error messages page 95 Cleaning and Maintaining the 1000 Thermal Cycler The 1000 thermal cycler requires little maintenance in order to maintain proper operation and precise thermal control However with long and constan
53. Supplies 0000 cee eae 58 Protocol Files and Folders 000 cece ene eens 61 Protocol File Contents 2 3 2 6 62 4 4 hota de Sa eo Ne ee ee eee 61 Protocol Steps 61 Options for Temperature and Gradient Steps 64 Managing protocols With the FILES Option 000 02 e eee 65 Copying Moving Deleting and Renaming Protocol Files 65 Copying a File With the copy Function 66 Moving a File With the MovE function 67 Deleting a File With the DELETE Function 68 Renaming a File With the RENAME Function 69 Protecting Files in a Folder With the SECURE Function 005 71 Creating Deleting and Renaming Folders 000000eneeeee 72 Creating a New Folder With the NEw Function 72 Deleting a Folder with the DELETE Function 73 Renaming a Folder With the RENAME Function 74 Viewing Protocol Files With the VIEW Option 0 000 eee 15 Advanced Tools and Functions 000200eenneeee 79 THe COL ODIONS caia aa aa a a 79 View the LAST RUN Tool 80 Test the Instrument With the SELFTEST Tool 81 Check the Firmware Version With the VERSION Tool 82 Name the Thermal Cycler with the NAME Tool 82 Change Default Parameters With the DEFAULTS Tool 83 View a Temperature Gradient with the GRADCALC Tool 84 Changing Screen View with the CONTRAST Tool 86 Change Settings With the PoRT Tool 87 Control an 1000 Thermal Cycler With the C1000 Thermal Cycler 88 Connecting Directly to a C1000 via the S1000 US
54. The C1000 thermal cycler will detect the S1000 thermal cycler 3 Connect additional 1000s directly to the C1000 Repeat steps 1 and 2 to connect up to three S1000s directly to the same C1000 4 Open the MAIN screen or instrument tree on the C1000 thermal cycler Open the MAIN screen or instrument tree on the C1000 thermal cycler or in the 1000 series software if the C1000 thermal cycler is connected to a computer 5 Select the S1000 thermal cycler Select the S1000 thermal cycler by serial number or name If the S1000 thermal cycler has a name page 82 then the name will be displayed instead of the serial number Connecting Indirectly to a C1000 Thermal Cycler via the 1000 USB A Port The S1000 thermal cycler can function under the control of the C1000 thermal cycler even when it is connected indirectly To accomplish this follow these instructions 1 Plug a shielded USB cable into the 1000 thermal cycler Plug a high quality shielded USB cable into the USB B port on the back of a 1000 thermal cycler page 8 A shielded USB cable can be ordered directly from Bio Rad 2 Plug the USB cable into the C1000 thermal cycler Plug the other side of the USB cable into a USB A port on the back of the C1000 The C1000 thermal cycler will detect the first S1000 thermal cycler 3 Connect another 1000 thermal cycler indirectly to the previous S1000 Plug a high quality shielded USB cable into the USB B port on the back of another
55. a Protocol File with the NEW Option 0000c eee eeee Adding INC EXT RATE and BEEP Parameters 000 cee eee eens Adding an Increment INC to a Temperature Step 28 Extending EXT the Hold Time in a Temperature Step 30 Changing the Ramp Rate RATE in a Temperature Step 31 Adding a Beep BEEP to a Temperature Step 32 Setting Temperature Control Mode 000c eee e eee e ene Choosing Lid Temperature 0002 cece ee ee eee eee Changing an Existing Protocol with the EDIT Option 005 Run Existing Protocols 000 cee ees Running a Protocol With the RUN Option 00 cece eee ees Monitoring a Run with the SCREEN Key 000 cee e eee eee Pausing and Resuming a Run with the PAUSE Key 000002 ees SKIDDING a SlCD eae yeni we wm be oe Ss eee ne hie eee eee Stopping a Run With the CANCEL Key 2 0 0 eee eee eee Incubating Samples With the INCUBATE Key 0c cece eee eee 1000 Thermal Cycler Manual Table of Contents Chapter 5 Chapter 6 Chapter 7 Optimizing PCR With the S1000 Thermal Cycler 53 Optimizing for Faster Protocols 00 cence e eee eee eee 53 Optimizing Temperature Steps With a Gradient 2000008 55 Optimizing Runs With Small Sample Volumes 0 00000005 56 Transferring Protocols From Another Thermal Cycler 5 57 Troubleshooting PCRs eee ee eee ee Se es 57 Microplate Tube Sealer and Cap
56. aling step Choose a gradient range that is as wide as possible to test for the optimal annealing temperature and bracket the calculated T by 5 to 12 C For example if the calculated T is 55 C then use a 20 C gradient to bracket that T In this example the gradient temperature range is from 45 to 65 C NOTE The widest possible gradient on the 1000 thermal cycler is 24 C 3 Choose a high annealing temperature from the results of the gradient PCR run By choosing a high annealing temperature you reduce the chance of nonspecific primer binding Select a high annealing temperature from the results of a PCR experiment with a gradient in the run See Step 2 on page 55 for details 55 1000 Thermal Cycler Manual Optimizing PCR With the S1000 Thermal Cycler about creating a gradient In this example the gradient ran from 55 to 67 C The best annealing is found at 60 C Temperature Temperature too low too high nonspecific bands faint bands Temperature good solid band and no nonspecific bands As a starting place use the highest annealing temperature and subtract 1 or 2 C Don t use the absolute highest temperature At higher temperatures the yield decreases resulting in faint bands Furthermore at high temperatures the primers might behave inconsistently from PCR run to PCR run 4 Narrow the temperature gradient in the annealing step optional Run a narrower annealing temperature gradient using the res
57. ample the file named STD3 is selected in the folder named EVA PROTOCOLS ITAQFHST Selected TOs Selected folder G file 3 Change the default lid temperature for this protocol optional Press the number keys to enter a new lid temperature Then press ENTER to continue to the next item In this example the lid temperature is changed to 110 C l Default lid File name A 3 re temperature PERT 3 55 0 4 m G A i Final hold c i 6 a lt temperature 34 Changing an Existing Protocol with the EDIT Option 4 Change the default sample volume optional Press the numeric keys to enter a sample volume Then press ENTER to continue to the next item NOTE Entering a sample volume between 1 and 50 selects Calculated mode standard Entering zero 0 selects Block mode For more information about these temperature control modes see page 33 In this example the oii volume is 20 ul and is unchanged Lg af as BLS ise te Lia FOR FOR FUR FOREVER mm ws ces TEA ai a A a T A ae Sj ao me el mat a IME el ed G 2 Se OF an TIP The sample volume can be also changed each time you run a protocol 5 Select a protocol step to edit Press the up or down arrow keys to select a protocol step Press ENTER to begin editing the step In this example step 4 is selected FOR FOR F OR reve te LT Pare b T T H E Selected step r ata m al T
58. ared it cannot be displayed again The total number of error messages that occur during a run is displayed on the LAST RUN screen page 80 However there is no process in the 1000 thermal cycler to open these error messages after they are closed at the end of a run If you want to track and log error messages run the 1000 thermal cycler under the control of a 95 1000 Thermal Cycler Manual Vaintenance amp Troubleshooting 96 C1000 thermal cycler or a robotic system that can retrieve a list of errors in the system and run logs Several error messages indicate problems that can be resolved or that might not change the results of your PCR The table below contains a list of error messages and possible solutions Table 14 Warning and error message solutions Message A C POWER FAILED POWER OUTAGE DURING CYCLE X STEP Y RESTARTED AT ZZ Z TO CONTINUE PRESS ENTER PLEASE RESTART CYCLER PLEASE CALL BIO RAD FOR SERVICE BLOCK OVERHEATED PLEASE RESTART CYCLER HEATSINK OVERHEATED PLEASE CALL BIO RAD FOR SERVICE PLEASE RESTART CYCLER SYSTEM OVERHEATED PLEASE CALL BIO RAD FOR SERVICE PLEASE CALL BIO RAD FOR SERVICE ALL BLOCK SENSORS FAILED PLEASE CALL BIO RAD FOR SERVICE POWER SUPPLY OVERHEATED PLEASE CALL BIO RAD FOR SERVICE HEATED LID FAILED PROTOCOL CANCELLED PLEASE CHECK AIRFLOW HTSINK OVERHEATING PLEASE CALL BIO RAD FOR SERVICE PLEASE CHECK AIRFLOW SYSTEM OVERHEATING PLEASE CALL BIO RAD F
59. as failed system using average of amplifier temperatures to continue 97 1000 Thermal Cycler Manual Vaintenance amp Troubleshooting Table 14 Warning and error message solutions Message PLEASE CALL BIO RAD FOR SERVICE LID OVERHEATED AND WAS SHUT OFF PLEASE CALL BIO RAD FOR SERVICE AMP1 TEMP SENSOR FAILED PLEASE CALL BIO RAD FOR SERVICE POWER SUPPLY SENSOR FAILED PLEASE CALL BIO RAD FOR SERVICE BLOCK POWER FAILURE PROTOCOL CANCELLED PLEASE CALL BIO RAD FOR SERVICE LOGIC POWER FAILURE PLEASE CALL BIO RAD FOR SERVICE BASE POWER FAILURE PROTOCOL CANCELLED PLEASE CALL BIO RAD FOR SERVICE AMP2 TEMP SENSOR FAILED BLOCK MISSING PROTOCOL CANCELLED PLEASE CALL BIO RAD FOR SERVICE PLEASE CALL BIO RAD FOR SERVICE MEMORY CORRUPT PROTOCOLS MAY BE LOST PLEASE RESTART CYCLER BAD REACTION MODULE PLEASE CALL BIO RAD FOR SERVICE PLEASE RESTART CYCLER INCORRECT CHECKSUM PLEASE CALL BIO RAD FOR SERVICE PLEASE RESTART CYCLER BLOCK POWER SHUT OFF PLEASE CALL BIO RAD FOR SERVICE PROTOCOL CANCELLED SAMPLES COOLED AT 4C BLOCK SENSOR FAILED AT CYCLE X STEP Y PLEASE RESTART CYCLER PLEASE CALL BIO RAD Cause duals only Lid has overheated and has been shut off and protocol has been terminated Amplifier temperature sensor 1 has failed Power Supply sensor has failed Power to block is out of range Logic power sensor is out of bounds Base power sensor is out of bounds Protocol cancelled
60. cludes descriptions and catalog numbers Table 24 Catalog numbers of accessories to the 1000 series number USB cable Shielded USB cable to connect the S1000 184 8000 thermal cycler directly to C1000 thermal cycler Tube Tray Provides structural support for one or a few 184 7000 tubes 112 Life Science Group 10010306 RevA US EG Bio Rad Laboratories Inc Web site www bio rad com USA 800 4BIORAD Australia 61 02 9914 2800 Austria 01 877 89 01 Belgium 09 385 55 11 Brazil 55 21 3237 9400 Canada 905 712 2771 China 86 21 6426 0808 Czech Republic 420 241 430 532 Denmark 44 52 10 00 Finland 09 804 22 00 France 01 47 95 69 65 Germany 089 318 840 Greece 30 210 777 4396 Hong Kong 852 2789 3300 Hungary 36 1 455 8800 India 91 124 4029300 Israel 03 963 6050 Italy 39 02 216091 Japan 03 5811 6270 Korea 82 2 3473 4460 Mexico 52 555 488 7670 The Netherlands 0318 540666 New Zealand 0508 805 500 Norway 23 38 41 30 Poland 48 22 331 99 99 Portugal 351 21 472 7700 Russia 7 495 721 14 04 Singapore 65 6415 3188 South Africa 27 861 246 723 Spain 34 91 590 5200 Sweden 08 555 12700 Switzerland 061 71795 55 Taiwan 886 2 2578 7189 United Kingdom 020 8328 2000 07 0535 1007 Sig 1106
61. col With the RUN Option The RUN option provides a quick way to choose and run an existing protocol Choose a protocol from the preinstalled protocols from the MAIN folder or any user folder in the file tree page 16 The following instructions assume you know how to operate the main menu and control panel page 13 To run a protocol follow these instructions 1 Select RUN in the main menu and load the samples in the block Check that the main menu status shows Block is idle then press the arrow keys to select RUN if it is not already selected Choose and select the protocol you want to run Finally press ENTER to confirm the selection and continue to the next screen Load samples into the block Then close the lid and set the lid force page 13 Use sufficient force on the lids to achieve the following e Push the tubes or microplate down into the block for complete contact with the block e Seal the sample lid to prevent evaporation of the sample 41 1000 Thermal Cycler Manual Run Existing Protocols In this example the RUN option is selected and the block is idle RUN option Status message i EF single block Bleck it idle NOTE With a dual 48 reaction module the status message is Blocks are idle when the blocks are both available to run a protocol 2 Select a protocol to run Press the arrow keys to select a folder and then press the right arrow key to begin selecting a file in that folder Choose the MAIN
62. d Edit Protocols Confirm the parameters The next screen displays three choices YES No and Option Select one of the choices by pressing the right and left arrow keys then press ENTER to continue to the next screen e Select YES to confirm the current parameters for this protocol step e Select No to change any parameters in this protocol step e Select Option to preview the temperature gradient PREVIEW described next or to add a hold time extension EXT described on page 30 In this example YES ts selected to confirm the current parameters Uol 26pL FUR FOR Three choices Ho Option Select Option to preview the gradient before the run optional In this example OPTION Is selected Hew STOSGRAD_ 45 a 7 Ho F T I OH Select PREVIEW to view the gradient In this example PREVIEW is selected Pa z T URTE Choose preview PREVI EHE Sea Press ENTER to preview the gradient press again to return to the previous screen 24 Creating a Protocol File with the NEw Option In this example the gradient is formed on a 96 well block with a range from 55 to 75 C This screen displays the approximate temperature of each row of the block and labels the front and back rows FROHT ROM Temperature in each row approximate Rows in block D h e O ey G iq in ty oy Ty lt Pe CT H Ei F E E F E H BACE ROM 7 Continue entering the protocol steps Repeat
63. d during the run e Firmware version on the S1000 thermal cycler during the run The TOOLS Options In this example the last protocol run is STD2 and the firmware version is 2 0 96 Protocol name r tad EL Turn of below temperature Lid temperature z VOLUME S l l ampe vo ume TOTAL TIME ERRORS VERSION Total run time Firmware version TIP If you want to run this protocol again note the name of the protocol then select the MAIN gt RUN and select the protocol by name Test the Instrument With the SELFTEST Tool Select the TOOLS gt SELFTEST tool to instruct the thermal cycler to run a test to check that it is running within specifications The S1000 thermal cycler automatically runs a self test every time it starts During the SELF TEST protocol the instrument checks that it is working correctly before a run Follow these instructions to manually run a self test 1 Select SELFTEST To select SELFTEST press the arrow keys Then press ENTER to continue to the next screen TOOLS LAST FUH Select SELFTEST SBELET EST WERSIOH HAME 2 View the self testing screen The thermal cycler displays the following self testing screen while it runs the tests You will also hear the instrument fans turn on and off during the test When the test has successfully completed the main menu displays SELF TEST IH PROGRESS gt E E eX 81 1000 Thermal Cycler Manual Advanced Tools and Functions
64. d turn on the thermal cycler Use the supplied power cord to plug the thermal cycler base into an electrical outlet Use the power switch on the back of the base to turn on the power NOTE Before operating the thermal cycler be sure to read the safety warnings and operating requirements listed in the Safety and Regulatory Requirements 1000 Thermal Cycler Manual Get Started As the instrument boots up the thermal cycler runs a self test to verify proper functions and then displays the main menu Use the main menu to begin operating the thermal cycler The main menu provides access to all thermal cycler operations and indicates cycler status If the thermal cycler is named the name displays in the top of the main menu For information about naming the thermal cycler see page 82 For example the main menu shows the options listed below the status message Blocks are idle and the thermal cycler name MENDEL Thermal cycler name Main menu options Status message lact are For more information about the options listed in the main menu see page 16 This thermal cycler and associated accessories are covered by a standard Bio Rad warranty Contact your local Bio Rad Laboratories office for the details of the warranty For more information about contacting your local Bio Rad Laboratories office see page 3 Thermal Cycler Overview Thermal Cycler Overview When complete the 1000 thermal cycler includes a base and
65. describes how to manage folders by creating deleting and renaming them In a folder the protocol files are listed in the order that they were created Therefore storing all files in the MAIN folder can result in a long unordered list of files making it difficult to find a file Alternatively storing protocol files in a user created folder has these advantages e Locate your protocol files easily without sorting through a long list of protocol files e Password protect your protocol files by storing them in a secure folder page 71 Secure folders prevent other instrument users from editing deleting or moving your files page 71 Securing protocol files in a folder does not prevent other instrument users from viewing running or copying the files TIP If you have not yet created a folder then you can temporarily store a new protocol file MAIN folder After you create your folder select MOVE page 67 to transfer your protocol files to that folder This section provides step by step instructions about managing folders e Select CREATE to make a new folder next section page 72 e Select DELETE to delete a folder page 73 e Select RENAME to change the name of an existing folder page 74 Creating a New Folder With the NEW Function Select FILES gt NEW to create a new folder when you want to store a set of protocol files in an easy to find location NOTE The 1000 thermal cycler can contain 11 folders in addition to the MAIN fo
66. dit Protocols 1 Select NEW to create a new protocol file Press the arrow keys to select NEW in the main menu Press ENTER to confirm the selection and continue to the next screen RUH WT EH New protocol HEH FILE EDIT TOOLS Blocks arg idle NOTE The 1000 thermal cycler s internal memory can hold up to 400 typical 2 STEP protocols 2 Name the new protocol file Enter a protocol name composed of letters and numbers To type a number press the numbered key To select a letter of the alphabet press the up or down arrow key Press ENTER to continue to the next space For example to select the letter c press the UP key 3 times To select the letter s press the DOWN key eight times In this example the default letter A is selected Enter a name here NOTE On the S1000 thermal cycler a protocol file name can contain from one to eight characters The characters are numbers or capital letters Each protocol name must be unique to all folders Enter the first letter In this example the letter s is entered Ltd T6576 Val i161 TIP For more information about how to enter a letter of the alphabet see Control Panel Keys page 15 20 Creating a Protocol File with the NEw Option In this example the characters STD3 are entered and the cursor is highlighting the next space Ready for next character TIP To delete or change a letter press CANCEL and select a new letter To delete the entire
67. duct Typically the cycle includes three temperature steps called denaturing annealing and extension steps These temperature steps denature the DNA template anneal the primer to the template and allow the polymerase to extend the complementary strand of DNA However under certain conditions the annealing and extension steps are combined in a shorter two step cycle To add the denaturing annealing or extension steps in the S1000 thermal cycler add a temperature step page 22 or gradient step page 23 To cycle these temperature steps add a GOTO step immediately after the last step that you want to repeat in the cycle page 25 A final temperature step to extend the PCR product This final extension temperature step serves to finish extending any PCR product that did not fully elongate during the cycles To add a final step in an S1000 thermal cycler protocol file add a temperature step page 26 Protocol steps on the S1000 thermal cycler vary from the typical PCR protocol steps When you transfer a typical PCR protocol to the 1000 thermal cycler you translate them into the four 1000 thermal cycler protocol steps e Temperature TEMP which is the same as a typical PCR protocol step e Gradient GRAD which is a temperature step that includes a gradient e Goto GOTO which is used to add repeats and create cycle in PCR e End END which instructs the thermal cycler to end the protocol The following is a composite scr
68. e power supply heat pump or other accessories If you open these instruments you put yourself at risk for electrical shock and void your warranty All repairs must be done by an authorized repair service Never remove the outer case of a thermal cycler base because you might receive an electrical shock This thermal cycler uses neutral fusing which means that live power could still be exposed inside the instrument even when the fuse is blown or removed Warning about risk of burning A thermal cycler generates enough heat to cause serious burns Wear safety goggles or other eye protection at all times during operation Always allow the sample block to return to idle temperate before opening the lid and removing samples Always allow maximum clearance to avoid accidental skin burns Table 2 Instrument safety warning labels Icon Meaning Warning about risk of explosion The sample blocks can become hot enough during the course of normal operation to cause liquids to boil and explode Safety Use Specifications and Compliance This instrument has been tested and found to be in compliance with all applicable requirements of the following safety and electromagnetic standards Table 3 Safe use specifications Safe use requirements Specifications Input power 100 240 VAC 50 60 Hz Ambient temperature of 5 C to 31 C Relative humidity Temperature Indoor use i maximum of 80 noncondensing Altitude Up to 2 000 meters above
69. e number of cycles in the protocol Before you adjust the number of cycles you must know the approximate concentration of PCR template To approximate the concentration of an unknown template start with 30 to 45 cycles the first time you run the protocol Then detect the PCR product in a gel stained with Ethidium Bromide and estimate the starting concentration If the concentration is sufficient then shorten the number of cycles by 5 and run the protocol again Once the concentration of the target sequence is known minimize the number of cycles in the protocol until the concentration of the PCR product is too low Choose the best number of GOTO repeats from the results of these reactions Optimizing Temperature Steps With a Gradient For an efficient reaction and clean product optimizing the annealing step is critical Use a gradient to optimize the temperature of the annealing step In addition use a gradient step to optimize other temperature steps Follow these instructions as a suggested approach to optimizing the annealing temperature 1 Calculate the predicted annealing temperature based on the melting temperature Tm of the primers and template Use a T calculator to find the T of any combination of primers and template When you run the 1000 thermal cycler under the control of a C1000 thermal cycler use the T Calculator available onboard the C1000 thermal cycler 2 Create a PCR protocol with a gradient step in the anne
70. e wells must be cleaned thoroughly and often Clean the oil when it is discolored or contains dirt Use a solution of 95 ethanol to clean oil Do not allow oil to build up in the block WARNING Never heat the block after adding a cleaning solution Heating the block with cleaning solution will damage the block lid and thermal cycler base e Clean the inner lid Use a soft cloth and water to remove debris and solutions from the inner lid surface Never use abrasive detergents or rough material that will scratch the surface Cleaning the inner lid improves precise sample heating and cooling e Clean the outer lid surface Use a damp cloth or tissue to clean spills off the outside case If needed use a mild soap solution and then rinse the surface with a damp cloth Cleaning the cover will prevent corrosion Maintaining Sufficient Air Flow The 1000 thermal cycler requires sufficient airflow to heat and cool precisely If the flow of air is blocked or too warm the thermal cycler cannot efficiently ramp to the correct temperature This section includes instructions for testing the air flow and suggestions for fixing low or warm air flow Testing for Sufficient Airflow Airflow is sufficient when the thermal cycler heats and cools to the correct target temperatures efficiently When you first set up the S1000 thermal cycler in a new location you can test for sufficient local airflow by following these instructions 1 Set up the instrume
71. een showing an S1000 thermal cycler protocol with all four types of protocol steps Hiei Protocol name 7 DAGRAD ki i j Temperature step includes a gradient step Ty cn Gradient step GOTO step 55 7 G 7 7 bol F Protocol File Contents Table 11 describes the four S1000 thermal cycler protocol steps with the name parameters limits and function Table 11 S1000 thermal cycler protocol steps and parameters Step Name TEMP temperature GRAD gradient GOTO END Parameters and Ranges TEMP C The target temperature between 0 0 and 100 0 C in tenths of a degree TIME The hold time between 1 second and 18 hours in the format of hours minutes seconds To enter an infinite hold Forever press the co Infinite 0 key Lower The lower temperature in the gradient Enter a number between 30 0 and 99 0 C in tenths of a degree Upper The upper temperature in the gradient The maximum temperature is 100 C Enter a temperature within 24 0 C of the Lower temperature TIME The hold time between 1 second and 18 hours in the format of hours minutes seconds To enter an infinite hold Forever press the co key zero 0 key GOTO STEP The step number of the first step in the repeat ADDTNL REPEATS The number of additional times that the steps repeat No parameters Description A temperature step that instructs the thermal cycler t
72. elect RUN if it is not already selected Then press ENTER to start running the protocol Rum ITAQFST Sample Hol 25eL Select RUN Fi UH WIT EH NOTE To run a PCR successfully a thermal cycler must be able to heat and cool quickly Do not block the air vents Keep other instruments and clutter away from the thermal cycler page 99 Monitor the protocol optional Once you start a run the Main Menu is displayed and the status line at the bottom of the screen will indicate that the protocol is running Pressing the lt Screen gt key displays the Running screen which shows the step parameters at the top of the list and the current status of the step below it In this example the lid is preheating before the first step of the protocol begins and the current lid temperature is 56 C Running Current step and parameter ITAHQFST C f ase z i Current sample p H 38 temperature oT i Preheating Lid 56 Current lid temperature Running a Protocol With the RUN Option Once the lid is heated to within 10 C of target temperature the S1000 thermal cycler begins the first step and first cycle of the protocol In this example the 1000 thermal cycler is heating to 98 C for Step 1 in the first cycle Running on A ITAQFST Target Step 1 a8 0t 0 temperature Temp Current cycle i seat 1 Current temperature TIP To have the instrument beep when the 1000 thermal cycler reaches the
73. emperature step in the protocol file Press the arrow keys to choose a step type Select TEMP to enter a temperature step To select GRAD and enter a gradient temperature step see Step 6 on page 23 for instructions Press ENTER to continue to the next screen In this example TEMP is selected rzit EHTER NOTE The first step in a protocol must be a TEMP or GRAD step Enter the target temperature between 0 0 zero and 100 0 C Press the number keys to type the target temperature for the temperature step Press ENTER to continue to the next item In this example the target temperature is 95 Target temperature Enter the hold time between 0 01 one second and 18 00 00 18 hours Enter 0 zero to add an infinite hold and hold this step FOREVER Press the number keys to type the hold time TIME in minutes and seconds For example to enter four minutes 4 00 type 400 or to enter 30 seconds type 30 Press ENTER to continue to the next field In this example the hold time is four minutes Hold time Confirm the parameters The next screen displays three choices YES NO and Option Select one of the choices by pressing the right and left arrow keys then press ENTER to continue to the next screen e Select YES to confirm the current parameters for this protocol step Creating a Protocol File with the NEw Option e Select No to change any parameters in this protocol step e Select Option to add more parameters to t
74. er through a USB cable then program and run protocols using the C1000 thermal cycler user interface e The 1000 series Manager software Connect the C1000 thermal cycler to a computer through a USB cable then control the C1000 thermal cycler and any connected 1000 thermal cyclers with the 1000 series software The following three sections describe how to connect and operate an 1000 thermal cycler using a C1000 cycler e Connect directly to a C1000 thermal cycler via the S1000 USB B port below e Connect indirectly to a C1000 thermal cycler via the S1000 USB A port page 89 e Operating the 1000 thermal cycler while under the control of the C1000 thermal cycler page 90 For detailed instructions about running the S1000 thermal cycler when it is connected to a C1000 thermal cycler see the instructions in the C1000 thermal cycler manual or the 1000 series software online help or manual Connecting Directly to a C1000 via the 1000 USB B Port To connect the S1000 thermal cycler directly to a C1000 thermal cycler follow these instructions Control an S1000 Thermal Cycler With the C1000 1 Plug a USB cable into the S1000 thermal cycler Plug a high quality shielded USB cable into the USB B port on the back of the 1000 thermal cycler page 8 For USB cable part numbers see page 111 2 Plug the USB cable into the C1000 thermal cycler Plug the other side of the USB cable into a USB A port on the back of the C1000 thermal cycler
75. ermal Cycler Optimizing a reaction is critical for successful and repeatable PCR Optimizing involves choosing the best reagents as well as choosing the most effective temperatures and hold times for each step of the protocol This chapter describes how to use the features of the 1000 thermal cycler to optimize PCR protocols and recommended PCR supplies e Optimize for a faster PCR protocol below e Optimize for efficient reactions by including a temperature gradient page 55 e Optimize runs with small sample volumes page 56 e Transfer protocols that were run on another thermal cycler page 57 e Troubleshooting PCR page 57 e Select compatible PCR supplies including microplates sealers tubes and caps page 58 Complete optimization of a protocol involves selecting appropriate reagents and enzymes as well as testing the parameters of the PCR protocol For more detailed information about optimizing protocols for fast PCR search the sites listed here These articles include tips for optimizing the reagents in addition to the tips for optimizing a PCR protocol presented in this section e Gene Expression Gateway bio rad com genomics Go to the web site and select Application Techniques gt Quantification gt Fast PCR e BioRadiations Magazine volume 118 in Adobe PDF format Go to discover bio rad com and search the Literature page 3 for Fast PCR Optimizing for Faster Protocols Optimizing a protocol for faster PCR
76. ess ENTER to continue to the next screen In this example the STD2G file is selected in the MAIN folder Dgelgte PROTOCOLS ho TEJ FLIG rotocol file amp a vH j GRAHT gt HHH a ae 40a D a m ohm oh ve Fan Pam mm l F NOTE If the protocol file is in a secure folder you must enter a password to delete a file 3 Confirm the deletion Select YES or No Select YES and press the ENTER key to delete the file and return to the main menu Select No to cancel the deletion WARNING Deleting a file permanently removes all records of the file Deleted protocol files cannot be retrieved Protocol names must be unique but once deleted a file name can be reused In this example DELETE is selected Select Delete pee Te Ce p A I I I I p Select YES Renaming a File With the RENAME Function Select FILES gt RENAME to rename an existing protocol file when you have edited the protocol steps and you want the name to reflect the change To rename a protocol file follow these instructions 69 1000 Thermal Cycler Manual Protocol Files and Folders 1 Select RENAME To select RENAME press the arrow keys Then press ENTER to confirm the selection and continue to the next screen FOLDERS HEH SECURE PELETE RENAME Select RENAME 2 Select a file to rename Press the up and down arrow keys to select a folder Then press the right arrow key to begin selecting a file Finally
77. f keys on control panel Keys Additional notes SCREEN Toggles between screens for Press this key to view the status of alternate views a run NUMERIC AND SYMBOL KEYS 1 through 9 Enter numbers Press each key to select numbers PAUSE Enter a decimal point or pause a protocol INCUBATE 0 x Enter a zero begin an instant incubation or enter an infinite hold Enter a minus sign or cancel a Press this key to delete an entry function cancel a function stop a protocol end an incubation or delete text on the screen CANCEL NAVIGATION KEYS RIGHT ARROW Move cursor to the right LEFT ARROW Move cursor to the left UP ARROW Move cursor up Press the up arrow key to scroll from A to z on the screen For example press the up key three times to select the letter c DOWN ARROW Move cursor down Press the down arrow key to scroll from z to A on the screen For example press the down arrow key eight times to select the letter s ENTER Confirm a selection a command an entry or to open a folder 15 1000 Thermal Cycler Manual Get Started The Main Menu The main menu provides access to all major functions that control the 1000 thermal cycler MEHODEL RUH LI EH Main menu __ HEH FILES options EDIT a es Blocks are idle TIP To access the main menu when the thermal cycler is not running a protocol press the CANCEL key several times until you exit the current function To access the main me
78. folder to select a preinstalled protocol Finally press ENTER to confirm the selection and continue to the next screen NOTE A protocol that is running cannot be edited Changes made in a protocol that is running apply to the next time the protocol runs In this example the preinstalled protocol ITAOFAST Is selected in the MAIN folder This protocol will run a fast PCR with iTaq polymerase PROTOCOL Selected protocol TIP You can run a protocol in a secure folder without the password See Protecting Files in a Folder With the SECURE Function on page 71 for more information about the files in secure folders 3 Select block Dual 48 module only Press the right and left arrow keys to select BLOCK AorBLOCK B Then press ENTER to continue to the next screen In a dual reaction module Block A is on the left side In this example BLOCK A is selected Stlect Black Block A selected BLOCK BLOCK E TIP When a block is selected the LED on the reaction module lights up 42 Running a Protocol With the RUN Option 4 Enter the sample volume The sample volume you enter determines the temperature control mode page 33 To use the Calculated mode standard enter the volume of your samples between 1 and 50 uL To use Block mode enter 0 zero NOTE Calculated mode is the recommended temperature control mode because it most accurately represents the actual sample temperature For more information see page
79. he arrow keys Then press ENTER to confirm the selection and move to the next screen FOLOERS PROTOCOLS HEH COPY SECURE MOVE ELETE DELETE Select RENAME BR FNAME REWHAME 14 Viewing Protocol Files With the VIEW Option 2 Select a folder to rename Press the up and down arrow keys to select the folder that you want to rename In this example the ANNA folder is selected Rename lt FOLDERS gt TILE ENA lt AHHA 3 Type the new name of the folder Press the up and down arrows to select letters and press the number keys to type numbers Then press ENTER to confirm a selection and move to the next space in the name When the name is complete press ENTER to confirm the name and return to the main menu NOTE A protocol file name can contain from one to eight characters The characters are numbers or capital letters Each protocol name must be unique NOTE To rename a folder that is secure enter the password before typing the name Viewing Protocol Files With the VIEW Option To view the content of protocol files and folders select the VIEW option in the main menu Follow these instructions to view files and folders 1 Select VIEW To select VIEW in the main menu press the arrow keys Then press ENTER to confirm the selection and continue to the next screen Select VIEW RUH LII EH HEH FILE EDIT TOOLS Blocks arg idle 2 Select a folder to view Press the up and down arrow keys to select
80. her e Shield the thermal cycler from heat sources such as radiators other heat producing instruments and bright sunlight Replacing Fuses The S1000 thermal cycler fuses are designed to blow in case of severe power surges or other sources of electrical short This process protects the user and instrument from excessive electric charge These fuses rarely need to be replaced However some institutions prefer to replace fuses on a regular basis to maintain full operability If the thermal cycler does not turn on first check that the power cord is plugged in to a functioning power source Additionally check that the power cord and power source are within the specifications for this instrument Table 21 on page 107 To replace a power cord contact Bio Rad Technical Support page 3 Finally check that the fuses are intact The S1000 thermal cycler runs with two fuses that have specifications listed in Table 21 on page 107 To remove and check the fuses follow these instructions WARNING Prevent electrical shock Always turn off and unplug the instrument from an electrical outlet before cleaning it 1 Use a small coin to unscrew the fuse drawer 2 Pull out the fuse drawer and remove the fuses 3 If a fuse is damaged replace it with a compatible fuse and close the drawer A bad fuse shows a break or burned spot in the metal A good fuse has intact metal Troubleshooting Troubleshooting The S1000 thermal cycler is des
81. his protocol step For more information about entering options see Adding INC EXT RATE and BEEP Parameters page 28 In this example YES is selected Hr Lid 116 C Wo 28eL Ho Option 6 Enter a gradient temperature step optional Press the right arrow key to select GRAD and enter a gradient temperature step Press ENTER to continue to the next screen NOTE A temperature gradient is limited to a 24 C spread The lowest possible Lower temperature in the gradient is 30 C and the highest Upper temperature is 100 C Therefore the lowest gradient is 30 54 C and the highest possible gradient is 76 100 C In this example GRAD Is selected Hori ST DSGRAD Choose gradient step TIP Check the temperature in each row of the block in a gradient by selecting the gradient calculator tool TOOLS gt GRADCALC on page 84 Enter the lower temperature in the gradient The lower temperature is at the front Row H of the block In this example the lower temperature is 50 C Lower Range of temperature temperatures back row to choose Enter the upper temperature in the gradient The upper temperature is at the back Row A of the block NOTE The range of temperature is limited by the widest available range for gradient which is 24 C The highest Upper temperature is 100 C Enter the hold time between 0 01 one second and 18 00 00 18 hours 23 1000 Thermal Cycler Manual Create an
82. igned to run without errors In general when the thermal cycler displays a warning or error message the message includes instructions for fixing the problem These sections discuss warnings and error messages e Restarting after a power failure below e 1000 thermal cycler error messages page 95 Restarting After a Power Failure If the external power is interrupted while the S1000 thermal cycler is running a protocol it resumes running the protocol as soon as power is restored In this case the following error message will alert the user that the protocol was interrupted Running om A Protocol name TTHOFA T FOUMER FAILED PUP E TEF 3 Details of the Hl oo Mon protocol step ee a el that was 1 COHTIWUE interrupted WARNING A power failure can change the outcome of a PCR run The hold time for the step that was running when the power failed is lengthened causing the sample to deviate from the target temperature until the power resumes Warning and Error Messages The S1000 thermal cycler tracks errors that occur during a run After a run all message display in sequence For example when a run is cancelled the following PROTOCOL CANCELLED screen displays Protocol cancellec na CAHCELLED screen Tofal Time Pal y Press the ENTER key to clear a message or error and continue to the next screen When all messages are cleared the S1000 thermal cycler returns to the main menu WARNING Once the message is cle
83. in the step Press the keys on the control panel to enter the temperature Then press ENTER to continue to the next parameter arrow keys In this example step 3 is selected Notice the original parameter for temperature is 55 0 C di 1 TO i el 4 HE 3z 5 ee H 38 Original j A H 3H parameters C i TI IH Choose EDIT In this example the temperature is changed from 55 to 65 C ott Wt oat ol a i te i LB Ga IJ i in Enter new parameter 1 Pe OF be od Pt oe Oe ee eH r k m ra Change the second parameter in the step Press the number keys to type a new parameter Then press ENTER to continue to continue to the next step In this example the hold time is changed to 35 seconds Enter a new parameter T ee maru m i a Zo E o a a Di a s T 1 4 38 Changing an Existing Protocol with the EDIT Option Continue editing the existing parameters in each step optional Press the up and down arrow keys to choose an earlier or later step in the protocol In this example the temperature parameter is changed to 70 0 C in step 4 a ea oa Lt A J pe E et a J 9 z lle at parameter in next step ye ee bi e a e a E ial 1 ma em p H es o a Press ENTER to finish changing parameters in the step and to continue editing the protocol In this example step 3 is finished and step 4 is selected Step 3 1a le
84. ion follow these recommendations e Place the thermal cycler base on a flat surface e Do not block fans during operation keep the area around the instrument clear of objects or debris that could block airflow e Read this manual for best usage practices e Never remove the instrument s outer casing e Do not operate in extreme humidity where condensation can impede proper operation e Do not operate at extreme altitude e Use only the provided power cord Table 21 Operating specifications Requirement Specification Thermal Block 0 4 C well to well uniformity at 90 C within 10 seconds Uniformity Thermal Accuracy 0 2 C of programmed target temperature Maximum Temperature 100 C Minimum Temperature 0 C 107 1000 User Manual Appendix B Operating Specifications Table 21 Operating specifications Requirement Max Ramp Rate Heated Lid Range Dimensions Weight Line Voltage Frequency Input Power Fuses Display Ports Specification 5 C second 0 110 C 13 x 18 x 8 23 Ibs 100 240 VAC rms no adjustment needed among voltages within these ranges 50 60 Hz single phase 400 W max Two 6 3 A 250 V 5 x 20 mm LCD semi graphical 5 USB A 1 USB B Gradient Specifications This thermal cycler can generate a temperature gradient across a 96 well or dual 48 well block To view the estimated temperature in each well use the GRADCALC tool 108 Table 22 Gradient specifications Requ
85. irement Gradient Direction Maximum Gradient Gradient Temperature Differential Gradient Uniformity Gradient Accuracy Gradient Calculator Accuracy Specification Back upper temperature to front lower temperature of block 30 C to 100 C 1 24 C 0 4 C well to well uniformity within 10 seconds of step Initiation 0 2 C of programmed target temperature 0 4 C of the actual well temperature 1000 Thermal Cycler Manual Appendix C Catalog Numbers This appendix lists useful catalog numbers and descriptions for the following components and accessories in the 1000 series e 1000 series components and descriptions below e Accessories for the 1000 series with descriptions Components of the 1000 series 109 1000 Thermal Cycler Manual Appendix C Catalog Numbers 110 Table 23 lists the components of the 1000 series instruments and software with the descriptions and catalog numbers Table 23 Catalog numbers for the 1000 series instruments and software number Thermal cycler bases and reaction modules C1000 thermal cycler Thermal cycler base for PCR and realtime 184 1000 base 1000 thermal cycler Thermal cycler base for PCR 184 2000 base 96 Fast reaction module 96 well block for PCR with adjustable 184 0096 heated lid Dual 48 Fast reaction Two 48 well blocks for PCR with adjust 184 0048 module able heated lid 384 reaction module 384 well block for conventional PCR with 184 0384 adju
86. itions have changed This increase could cause some secondary PCR products but should also increase the primary product Adjust the temperature in each step to reoptimize the protocol Most importantly reoptimize the annealing temperature page 55 Troubleshooting PCR This section is a quick guide for PCR troubleshooting options For more detailed and extensive troubleshooting go to the PCR Doctor To find the PCR Doctor open the Gene Expression Gateway bio rad com genomics in a web browser window and select Support gt Amplification Central gt PCR Doctor Follow these suggestions to re optimize a reaction that fails by adjusting the protocol Nonspecific PCR products in addition to the target product Nonspecific products result from mispriming When mispriming is not a result of a reagent problem then adjust the annealing temperature Increase the annealing temperature to increase specificity of primer binding To find the optimal annealing temperature use a gradient page 23 Nonspecific PCR products without the target product Nonspecific product with no target production is a result of complete mispriming When the mispriming is not a result of a reagent problem then adjust the hold time Increase the hold time in the annealing and extension steps to increase specificity of primer binding provide more time for complete extension Low yield of the target PCR product Low PCR yield is a result of mispriming an over
87. lder To create a new folder follow these instructions Creating Deleting and Renaming Folders 1 Select NEW NEW is the default selection when you first open the FILES menu If NEW is not selected press the arrow keys to select it Then press ENTER to confirm the selection and continue to the next screen PROTOCOLS Select NEW cory MOVE DELETE RENAME NOTE The 1000 thermal cycler can contain up to 12 folders including the MAIN folder 2 Type the folder name Press the up and down arrows on the keypad to select a letter and press the number keys to type a number Press ENTER to accept a selection and move to the next space When you complete the name press ENTER to accept the name and return to the main menu NOTE A folder name can contain from one to eight characters The characters are numbers or capital letters Each folder name must be unique to the 1000 thermal cycler In this the folder name of the new folder is ANNA FOLDERS PROTOCOLS cory SECURE MOLE DELETE DELETE REHANMNE REHAME Enter a new folder name 9 45ie aa E gt Lae TIP To secure the new folder and all files in that folder follow the instructions on page 71 Deleting a Folder with the DELETE Function Select FILES gt DELETE to completely remove an existing folder when it is no longer needed Alternatively rename the folder to continue using it for a different purpose page 74 WARNING Deleting a folder permanently rem
88. local Bio Rad Laboratories office page 3 or online discover bio rad com NOTE Sealing wax such as Chill out is recommended to seal small sample volumes of less than 10 uL Wax solidifies at room temperature Pierce the solid wax with a micropipet tip to remove the sample For more information about optimizing protocols for small sample volumes see Optimizing Runs With Small Sample Volumes page 56 Table 10 lists the tested sealing options for microplates and tubes offered by Bio Rad However if you have a verified source of these supplies the S1000 thermal cycler is compatible with a wide variety of PCR supplies Find a full list of supplies for PCR in any of these locations e Life Science Research Product catalog See Bio Rad Resources and References on page 3 for information about how to order a copy e Bio Rad Life Science website discover bio rad com Go to the web site and select Products gt Amplification e Gene Expression Gateway bio rad com genomics Open the website and select Catalog Index gt Amplification In Table 10 each series of supplies is listed with the prefix of the catalog numbers Table 10 Compatible sealing options for PCR microplates and tubes TUBES AND MICROPLATES CAPS AND SEALERS Description Description Catalog Prefix Wel ane NUMBER Catalog Prefix Tubes Caps TCS Individual tubes with caps TFI and TWI Strip tubes TBS 0 2 ml 8 and 12 tube P Strip
89. ly short extension hold time or too high an annealing temperature When the low yield is not a result of reagent problems adjust the protocol Run a touchdown protocol page 102 to increase amplification of the target product Alternatively decrease the annealing temperature or run a gradient to optimize the annealing temperature page 55 57 1000 Thermal Cycler Manual Optimizing PCR With the S1000 Thermal Cycler Microplate Tube Sealer and Cap Supplies 58 The 1000 series thermal cyclers can run using a large variety of standard supplies such as microplates tubes sealers and caps The supplies listed in this section are a subset of supplies that are verified by testing They function within the specifications of the 1000 thermal cycler thermal cycler However the S1000 thermal cycler is designed to work well with a broad range of standard supplies TIP The composition and thickness of these supplies influences the outcome of a reaction Whenever you significantly change the source or composition of supplies it is good practice to reoptimize the protocol before running an important experiment Microplates tubes sealers and caps come in a variety of compositions and colors Bio Rad recommends the standard supplies listed in Table 10 for compatibility with the 1000 series thermal cyclers For a full list of available reagents and supplies refer to the Life Sciences catalog which is available in print by contacting your
90. mal cycler returns to the main menu press the SCREEN key to view the LAST RUN screen This screen displays a synopsis of the last protocol that was run To return to the main menu press the SCREEN key again NOTE For Dual 48 blocks press the SCREEN key again to view the LAST RUN screen for each block The LAST RUN screen provides useful information to use for troubleshooting PCR experiments For information about optimizing your PCR runs with the 1000 thermal cycler see page 53 In this example the ITAQFST protocol was the last run LAST RUH Lid temperature jibe eile Sample volume fhi Al gla Total run timed E ss ae Errors that occurre 3i J Tri E during the run j Firmware used during run ot TIP The LAST RUN screen is also available in the TOOLS option page 80 10 Press the SCREEN key to return to the main menu Monitoring a Run with the SCREEN Key Once a protocol is running you can monitor the progress of the run with any one of three screens Press the SCREEN key to toggle through the three screens TIP For the Dual 48 reaction module the thermal cycler displays these screens for Block A and then for Block B Pressing the SCREEN key toggles the display through each screen and both blocks Each time you press the SCREEN key the instrument shows the following three screens in this order starting from the main menu 1 The Running screen This screen displays the current step parameters temperature
91. n Module C1000 48 48 System 185 1048 C1000 chassis Dual 48 48 Rxn Module C1000 384 well System 185 1384 C1000 chassis 384 well Rxn Module S1000 Fast 96 well System 185 2096 S1000 chassis Fast 96 well Rxn Module S1000 48 48 System 185 2048 1000 chassis Dual 48 48 Rxn Module S1000 384 well System 185 2384 S1000 chassis 384 well Rxn Module 1000 Thermal Cycler 184 2000 1000 thermal cycler chassis only 96 well Fast Reaction Module 184 0096 96 well Fast Rxn Module for 1000 Series 1000 Thermal Cycler Manual ntroduction Table 4 1000 Series Catalog Numbers Product Catalog Number Item Description 48 48 well Fast Reaction 184 0048 Dual 48 48 Rxn Module for 1000 Series Module 384 well Reaction Module 184 0384 384 well Rxn Module for 1000 Series CFX96 Real Time System 185 5096 C1000 chassis CFX96 Rxn Module CFX384 Real Time System 185 5384 C1000 chassis CFX384 Rxn Module CFX Manager Software 184 5000 PC Software for use with Real Time PCR CFX Software Security Edition 184 5001 Single User License real time software 1000 Thermal Cycler Performance and Features The S1000 thermal cycler features superb thermal performance with the flexibility to expand throughput It can perform standard polymerase chain reactions PCR cycle sequencing or other techniques that require controlled temperature incubation The S1000 operates as a stand alone thermal cycler with the following features e Reduced Protocol Run Times The
92. name press CANCEL multiple times When the protocol name is complete press ENTER to continue to the next screen Protocol name complete 3 Enter a lid temperature optional Press the numeric keys to type a new lid temperature Press ENTER to accept the lid temperature and continue to the next screen The lid temperature can range from 0 to 110 C The default S1000 thermal cycler lid temperature for new protocols is 105 C NOTE When the block is running an infinite hold at a temperature below the Turn off below parameter the lid heater maintains 31 0 C To change the default Turn off below parameter select TOOLS gt DEFAULTS page 83 In this example the lid temperature is changed to 110 C Lid temperature Heu Lid EEE C for new protocol STDZGRAD Ua Loy 4 Enter a sample volume in microliters uL Press the number keys to type the volume of your sample Press ENTER to continue to the next screen The default sample volume for a new protocol is 10uL NOTE Entering a sample volume between 1 and 50 selects Calculated temperature Control mode standard Entering zero 0 selects Block 21 1000 Thermal Cycler Manual Create and Edit Protocols 22 mode Calculated mode is the recommended mode because it most accurately represents the actual sample temperature For more information about Temperature control modes see page 33 TIP The sample volume can be changed each time you run a protocol Enter a t
93. nd the temperature is currently ramping up to the target temperature Running on A ITAQFST Step 1 9 8 6 8 36 Te t Bek i Teme ao St PAUSE PAUSE Protocol to the target ibd ALE paused temperature 48 Skipping a Step Once the protocol reaches the target temperature it maintains that temperature throughout the pause Protocol Repeat eee 2 To resume the protocol press PAUSE again Press the PAUSE key again while on the Running Screen or Graphical Screen to Instruct the thermal cycler to continue running the protocol In this example step 1 continues and the hold time begins counting up from 0 to 30 seconds Running ITAQFET i oe aS eS eS Temp g5 0 8 8l Hold time Cycle 1 counts up Skipping a Step If you need to skip a step in a protocol that is running press the ENTER key when the step you want to skip is running The thermal cycler will skip to the next step in the protocol TIP The S100 cannot skip a GOTO repeat unless the 1000 is controlled by a C1000 thermal cycler To skip a protocol step follow these instructions 1 Press ENTER while the step you want to skip is running Press the ENTER key to instruct the thermal cycler to skip the current step in the protocol Then press the right and left arrow key to select Yes or No Finally press ENTER again to confirm your selection 2 Notice that the next step is now running In this example step 3 is running in Cycle 4 Running
94. nical notes manuals product information and technical support press the following links to download or request a copy of this manual or other Bio Rad Laboratories literature e press the PDF icon to download a portable document format copy and open it using Adobe Acrobat Reader software www adobe com e press the folder icon and order a printed copy e press the FAX icon to request a FAX copy e Phone your local Bio Rad Laboratories office to request a printed copy In the United States and Canada call 1 800 424 6723 toll free phone and select the Literature option Use the following resources to locate what you need Table 5 Bio Rad resources Resource Local Bio Rad Laboratories representatives Technical notes and literature Technical specialists How to contact Find local information and contacts on the Bio Rad Laboratories web site by selecting your country on the home page bio rad com Also find the nearest international office listed on the back of this manual Go to the Gene Expression Gateway bio rad com genomics and locate the Search box in the upper right corner of the web page Type a search term in the box to find links to products technical notes and manuals Bio Rad Laboratories provides quality technical support We staff our Technical Support department with experienced scientists to provide our customers with practical and expert solutions To find technical support on the web go to the
95. nt where you plan to use it and turn on the power For instructions about setting up the thermal cycler see page 5 2 Adjust the local environment for typical conditions Turn on any nearby equipment such as fans Also open any window blinds to achieve optimal conditions during a run If more than one thermal cycler is in the area run a protocol on all the thermal cyclers at the same time 3 Runa typical PCR protocol for 30 minutes You do not need to include a sample but must include an empty microplate or tubes The lid will not heat correctly if it touches the hot block of the reaction module 4 Measure the air temperature at the air intake vents If the air intake temperature increases above 31 C use the following information to ensure sufficient airflow 93 1000 Thermal Cycler Manual Vaintenance amp Troubleshooting 94 Fixing Insufficient Airflow If the air temperature near the thermal cycler is above 31 C make one or more of the following changes to increase and optimize the airflow around a thermal cycler e Increase circulation of the air in the room by adding fans or adjusting the air conditioning system e Adjust air conditioning to lower the ambient air temperature e Move the thermal cycler to another location e Provide more space around the S1000 thermal cycler and between adjacent instruments Arrange instruments so that the warm exhaust of one instrument does not enter the air intake vents of anot
96. nu when a protocol is running press the SCREEN key to toggle through a set of screens including the main menu Select the options in the main menu to start these instrument functions RUN Select the RUN option to choose an existing protocol file to run NEW Select the NEW option to create a new protocol file EDIT Select the EDIT option to modify stored protocol files FILES Select the FILES option to copy move rename delete or secure protocol files and or folders VIEW Select the VIEW option to review an existing protocol file TOOLS Select the TOOLS option to change thermal cycler settings or to view the last protocol that was run The File Tree 16 All protocol files are stored in the file tree The file tree displays when you need to select a protocol to run select a protocol to edit or select a protocol to view For example when you select RUN in the main menu the file tree opens to allow you to select a file to run The file tree includes these folders e MAIN folder This folder stores the preinstalled protocols This folder cannot be deleted or renamed This folder can also store user created files but this practice is not recommended NOTE Protocol files stored in the MAIN folder are not secure e User folders These folders are created by S1000 thermal cycler users for storing user created protocol files User folders and all the files inside them can be secured with a password The following image shows an
97. nue to the next screen Step 3 A D i TELE j T2 8 A 38 IT 30 TIF Select B T2 0 a YES or No mi aes 11 Finish editing the protocol To finish editing select the final step in a protocol the END step Press the down arrow key to select the END step number Then press ENTER to continue to the next screen Choose END to finish the protocol Press the left arrow key to select END Then press ENTER to finish editing the protocol In this example END is selected Select END to finish editing IHEERT 40 1000 Thermal Cycler Manual 4 Run Existing Protocols The S1000 thermal cycler runs protocols intuitively Select the RUN option in the main menu and choose a protocol file to run For your convenience use the preinstalled protocols to quickly run a standard protocol This chapter provides instructions about how to run a protocol and using the INCUBATE key to incubate samples at a single temperature Run and monitor a protocol below Monitor a protocol by pressing the SCREEN key page 46 Pause and resume a protocol by pressing the PAUSE key page 48 Skipping a step during a run page 49 Stop a protocol by pressing the CANCEL key page 50 Incubating samples at a single temperature by pressing the INCUBATE key to run an instant incubation page 50 The instructions in this chapter assume you know how to use the control panel to operate the 1000 thermal cycler page 13 Running a Proto
98. o ramp to the specified target temperature and hold that temperature for the specified amount of time see Step 5 on page 22 A temperature step that instructs the thermal cycler to ramp to the specified target temperature gradient across the block and hold that temperature gradient for the specified amount of time see Step 6 on page 23 A protocol step that instructs the thermal cycler to repeat a set of steps for the specified number of times see Step 8 on page 25 NOTE The total number of cycles in the protocol is the number of GOTO repeats plus the first cycle A protocol step that instructs the thermal cycler to finish the protocol see Step 10 on page 27 63 1000 Thermal Cycler Manual Protocol Files and Folders 64 Options for Temperature and Gradient Steps Each 1000 thermal cycler temperature or gradient step can include options Most of the options add parameters to the protocol step while creating or editing the protocol steps The following shows the four options that apply to temperature steps ExT RATE BEEP The following shows the two options that apply to gradient steps Only the EXT option modifies the parameters in a gradient step PREVI EWES 4 Table 12 describes all five options for temperature and gradient steps and includes the limits and range of the parameters Table 12 Options for temperature and gradient steps Option Parameter and Range Description INC A temperature from
99. oelectrique edicte par le Ministere des Communications du Canada Cables Shielded cables must be used with this unit to ensure compliance with the Class A FCC limits Use only Bio Rad USB cable Catalog No 184 8000 with your 1000 Series cyclers 1000 Thermal Cycler Manual Safety and Regulatory Compliance Table of Contents Chapter 1 Chapter 2 Chapter 3 Chapter 4 IEEE OGUIGUOUN sisi ie sece Scere osm Goa eee Sree ce iene ee eee The Bio Rad 1000 Series Platform 0 0000 e eee es 1000 Thermal Cycler Performance and Features 000000uuee Bio Rad Resources and ReferencesS 0000 cee n eee nee ees Writing Conventions Used in This Manual 000000eeeeuee Get Started 5566 25 ee cee Gare nea a e A Setting Up the S1000 Thermal Cycler 0000 cee ees Thermal Cycler Overview sere S624 cote t ieee ses shee es eta kew eee Front View without Reaction Module 7 Top View with Reaction Module 7 Back View with Reaction Module 8 Interchangeable Reaction Modules 0000n eee nen nee neenee Inserting and Removing a Reaction Module 10 Opening and Closing the Adjustable Lid 20 0000 eee eee Opening the Lid 12 Closing the Lid and Setting Lid Force 13 Control Panel Keys and Main Menu 0000 cee ence eeneneee Control Panel Keys 15 The Main Menu 16 MING FIG SINC Co io eee cee hoe ere ees we te Sk a ee et ate ee ee Create and Edit Protocols 00 ccc eee ees Creating
100. oth Remove any heavy dust that is deep in the vents with a vacuum cleaner Use water and a soft cloth to remove debris that is stuck to the fins Avoid scratching the surface Never use cleaning solutions that are corrosive to aluminum such as bleach or abrasive cleansers If needed use a mild soap solution and rinse well to remove residue completely Cleaning the fins improves precise sample heating and cooling WARNING Clean the block Use a soft cloth and water to remove debris from the outer block Never use abrasive detergents caustic solutions or rough material that will scratch the block WARNING Never clean the block with strong alkaline solutions strong soap ammonia or high concentration bleach Never use corrosive or abrasive cleaning solutions These cleaning agents can damage the block and prevent precise thermal control e Clean the block wells Clean spills immediately to prevent them from drying Use disposable plastic pipettes with water recommended 95 ethanol or a 1 100 dilution of bleach in water Always rinse the wells with water several times to remove all traces of ethanol bleach or soap Maintaining Sufficient Air Flow WARNING Bleach ethanol or soap that is left in the blocks could corrode the block and or destroy tubes and microplates during a run Always rinse the block well after cleaning it with any solution other than water Use of oil in the wells is not recommended However if oil is used th
101. otocols optimized to run with iTaq hot start polymerase page 102 iProof protocols optimized to run with iProof high fidelity polymerase page 103 Reverse transcription protocols to amplify DNA from an RNA template page 104 Nested primer protocols to amplify a specific DNA sequence from a large complex DNA template page 104 Standard Protocols Two standard protocols that run two step and three step PCR with a standard DNA polymerase Choose these protocols to begin running PCR with new primers and DNA template or copy them to begin writing a new protocol Table 15 lists the parameters in each protocol Table 15 Standard Protocol STD2 STD3 To Run a Standard Two Step To Run a Standard Three Step Protocol Protocol Target Target Temperature C e E Temperature C Time min sec or Repeats or GOTO Step or GOTO Step 101 1000 Thermal Cycler Manual Appendix A Preinstalled Protocols Table 15 Standard Protocol Touchdown Protocol A touchdown protocol tests for the best annealing temperature for a specific primer template pair Choose this protocol to test a new set of primers and DNA template for the optimal annealing temperature Table 16 lists the parameters in each protocol Table 16 Touchdown Protocol To run a protocol with an increment temperature step that automatically finds the most stringent conditions for primer binding Target Temperature C a or GOTO Step Time min
102. oves it NOTE A folder that contains protocols will not be deleted Before you delete a folder select VIEW in the main menu to view the contents of the folder page 61 To delete a folder first empty the folder by moving or deleting all protocol files page 65 To delete a folder follow these instructions 73 1000 Thermal Cycler Manual Protocol Files and Folders 1 Select DELETE To select DELETE press the arrow keys Then press ENTER to confirm the selection FOLDERS PROTOCOLS NEW COPY SECURE MOWE Select DELETE DELETE NELETE REHAME REHAME 2 Select a folder to delete Press the up and down arrow keys to select a folder Then press ENTER to continue to the next screen In this example the ANNA folder is selected Oelete Folder lt MAIH gt lt EWH gt Select a folder z AHHA to delete TIP If the folder is secure you must enter the password to delete it 3 Confirm the deletion Press the arrow keys to select YES or No Select YES to delete the folder or select No to cancel the deletion Then press ENTER to confirm your selection and return to the main menu NOTE A folder that contains protocol files will not be deleted Renaming a Folder With the RENAME Function Select FILES gt RENAME to rename an existing folder Renaming a folder does not change the protocol files stored in that folder To rename a folder follow these instructions 1 Select RENAME To select RENAME press t
103. peratures to the back Row A hottest temperatures of the block Follow these instructions to view a gradient 84 The TOOLS Options 1 Select GRADCALC To select GRADCALC press the arrow keys Then press ENTER to continue to the next screen 45 aoe Select GRADCALC tAOCALE TRAST T 2 Enter the lower temperature in the gradient Press the number keys to type the lower gradient for the back row of the block Press the CANCEL key to type a decimal point The limit is 30 0 to 99 0 degrees In this example the lower temperature is 65 C Lower temperatur in gradient Limits for lower BS ae temperature 3 Enter the upper temperature in the gradient Press the number keys to type the upper gradient for the back row of the block Press the CANCEL key to type a decimal point The upper temperature must be greater than the lower temperature and be within 24 0 C of the lower temperature In this example the upper temperature is 75 C LOWER TEMP C Upper temperature in gradient Limit for upper C temperature NOTE The upper temperature is limited to a temperature that is within the widest possible gradient The widest possible gradient is 24 C 4 View the temperature gradient The temperature gradient is displayed as the temperature in each row of wells in the block 85 1000 Thermal Cycler Manual Advanced Tools and Functions 86 In this example the temperature in row D of this 96
104. pt Protocol Target Temperature C or GOTO Time min sec or Nested Primer Protocols Nested primers amplify a specific DNA sequence from a large complex DNA template such as genomic DNA Table 20 lists the parameters in each protocol Table 20 Nested Primer Protocol NESTPR2 NESTPR3 To Run Two step Protocol Using To Run a Three step Protocol Using Nested Primers Nested Primers Time min sec or Repeats Time min sec o or Repeats Temperature C or GOTO Step a a 700 eomz 3x Infinite hold CO 5 72 7 00 Infinite hold GOTO 8 CO 104 Nested Primer Protocols Table 20 Nested Primer Protocol Infinite hold OO 105 1000 Thermal Cycler Manual Appendix A Preinstalled Protocols 106 S1000 User Manual Appendix B Operating Specifications The S1000 thermal cycler is designed for repeatable consistent results It heats and cools very fast during operation Users should follow the operating specifications listed in this section to achieve reliable results e Instrument operating specifications and recommendations e Gradient specification Instrument Operating Specifications This thermal cycler is designed to operate safely in a standard research laboratory for research applications NOTE This instrument is for research use only For safe and reliable results the operation of this instrument must follow the specifications listed in this manual To ensure safe operat
105. rate Press the number keys to type the parameter in C second Press the PAUSE key to enter a decimal point NOTE Enter a ramp rate between 0 1 and 5 C second in tenths of a degree 31 1000 Thermal Cycler Manual Create and Edit Protocols In this example the rate is 2 0 The thermal cycler will ramp to the target temperature for step 3 at 2 0 C second z H Enter the rate parameter 4 Confirm the step parameters Press the arrow keys to select YES or No Select YES to confirm the ramp rate and finish entering the parameters for step 3 Select No and change the parameters for this protocol step Adding a Beep BEEP to a Temperature Step The BEEP parameter instructs the thermal cycler to emit a sound when the temperature reaches its target A BEEP can be added to any temperature step TIP Add the beep step to a temperature step such as an infinite FOREVER hold when you want the thermal cycler to signal that it has initiated the step To add a beep follow these instructions 1 Choose OPTION after entering the initial temperature step parameters Press the right and left arrow keys to choose OPTION Then press ENTER to continue to the next screen In this example the OPTION will be added to step 7 parameters FORE WE R Heo F T I OH 2 Choose BEEP to signal the end of the protocol step Press the right and left arrow keys to choose the BEEP parameter 32 Setting Temperature Control Mode
106. re step can contain up to four additional parameters Add these parameters by selecting OPTION Step 5 on page 22 Table 9 describes the parameters and where to find instructions about adding them to protocol steps Table 9 Additional parameters for temperature steps name Increment page 28 Changes the target The range is 10 to 10 C cycle in INC temperature of a step tenths of a degree within the by the specified temperature limits of a temperature increment each cycle step 0 100 C A step with an increment must be within a GOTO repeat in order to increment the temperature with each cycle Extension page 30 Extends the hold time The range is 60 to 60 seconds cycle EXT by the specified in whole numbers within the time number of seconds limits of a temperature step 1 each cycle second to 18 hours A step with an extend must be within a GOTO repeat in order to extend the step with each cycle Ramp rate page 31 Changes the ramping The range is 0 1 to 5 C second in RATE rate tenths of a degree for temperature steps Ramp rate can only be changed on the dual 48 and 96 well reaction modules The default ramp rate is the thermal cycler s maximum rate for the reaction module see Interchangeable Reaction Modules on page 9 Beep page 32 Instructs thermal No range Only applies to BEEP cycler to emit a temperature steps sound when it reaches the target temperature in a temperature step
107. return to the TOOLS options If you want to return to the original contrast press the CANCEL key to reject the changes Change Settings With the PORT Tool When you set the S1000 thermal cycler for remote control select the port that you want to use for data transfer with the PORT tool Select TOOLS gt PORT tool to instruct the thermal cycler to list the data transfer ports page 8 To select a port follow these instructions 1 Select PORT tool To select PORT press the arrow keys Then press ENTER to continue to the next screen TOO LA DEFAULTS a all GRADCALC WIE COUHT RAST Select PORT HA PORT 87 1000 Thermal Cycler Manual Advanced Tools and Functions 2 Select a port Press the up and down arrow keys to select a port from the list of all ports on the thermal cycler Then press ENTER to return to the TOOLS option SELECT PORT Select a port 3057606 Control an 1000 Thermal Cycler With the C1000 Thermal Cycler 88 Up to three S1000 thermal cyclers can run under the control of a C1000 thermal cycler The S1000 thermal cycler s can connect directly to the C1000 or in a daisy chain configuration where the S1000 thermal cycler is connected in series to a C1000 thermal cycler When connected to a C1000 thermal cycler the 1000 thermal cycler can be controlled by The C1000 thermal cycler control panel Connect the 1000 thermal cycler directly or indirectly to the C1000 thermal cycl
108. rongly recommend that you follow the safety specifications listed in this section and throughout this manual Safety Warning Labels Warning labels posted on the instrument and in this manual warn you about sources of injury or harm Refer to Table 1 to review the meaning of each safety warning label Table 1 Meaning of safety warning label CAUTION Risk of danger This symbol identifies components that pose a risk of personal injury or damage to the instrument if improperly handled Wherever this symbol appears consult the manual for further information before proceeding CAUTION Risk of electrical shock This symbol identifies components that pose a risk of electrical shock if improperly handled CAUTION Hot surface This symbol identifies components that pose a risk of personal injury due to excessive heat if improperly handled Instrument Safety Warnings The following warnings labels display on the instrument and refer directly to the safe use of this S1000 thermal cycler Table 2 Instrument safety warning labels Icon gt e BE Meaning Warning about risk of harm to body or equipment Operating the S1000 thermal cycler instrument before reading this manual can constitute a personal injury hazard Only qualified laboratory personnel should operate this instrument Warning about risk of harm to body or equipment from electrical shock Do not attempt to repair or remove the outer case of this thermal cycler bas
109. s the INCUBATE 0 key Once the incubation is started it continues until stopped To stop the incubation press the CANCEL key WARNING Do not incubate samples overnight at a high temperature Heating tubes or microplates indefinitely at a high temperature can evaporate all the liquid in your samples To incubate samples follow these instructions 50 Incubating Samples With the INCUBATE Key 1 Load your samples into the thermal cycler block and press INCUBATE Press the INCUBATE key to begin an incubation after loading the samples NOTE For the Dual 48 reaction module you must select the block that contains your samples before continuing to the next screen 2 Enter a lid temperature Select YES or No To heat the lid during the incubation select YES and press ENTER To turn off the lid during incubation select No and press ENTER In this example YES is selected Ae ee es ee Select YES or No TEBE Enter the lid temperature if you want to use the heated lid Press ENTER to accept the default lid temperature Press the number keys to type a new lid temperature In this example the lid temperature is 100 C Lid temperature es ee ie oe De Oa aa A L a 3 Enter an incubation temperature Press ENTER to accept the default incubation temperature of 75 C Press the number keys to type a new incubation temperature between 0 0 and 100 0 C Press the PAUSE key to enter a decimal point To delete a number
110. sea level Overvoltage Categories II Pollution degree 2 SAFETY COMPLIANCE This instrument has been tested and found to be in compliance with all applicable requirements of the following safety and electromagnetic standards e UL Std No 61010A 1 Electrical Equipment for Measurement Control and Laboratory Use Part 1 General Requirements e UL Std No 61010A 2 010 Electrical Equipment for Measurement Control and Laboratory Use Part 1 General Requirements e CAN CSA C22 2 No 1010 1 92 Safety Requirements for Electrical Equipment for Measurement Control and Laboratory Use Part 1 General Requirements includes Amendment 1 e CAN CSA C22 2 No 1010 1B 97 Amendment 2 CAN CSA C22 2 No 1010 1 92 Safety Requirements for Electrical Equipment for Measurement Control and Laboratory Use Part 1 General Requirements e CAN CSA C22 2 No 1010 2 010A 97 Safety Requirements for Electrical Equipment for Measurement Control and Laboratory Use Part 2 010 Particular Requirements for Laboratory Equipment for the Heating of Materials Amendment No 1 e EC 61010 1 Safety Requirements for Electrical Equipment for Measurement Control and Laboratory Use Part 2 010 Particular Requirements for Laboratory Equipment for the Heating of Materials ELECTROMAGNETIC COMPATIBILITY EMC e F C C Title 47 Part 15B as a Class A digital device EN61326 Class A Electrical Equipment for measurement control and laboratory use EMC Requirements
111. stable heated lid C1000 96 well Fast Sys C1000 chassis and 96 Fast reaction mod 185 1096 tem ule system C1000 48 48 Fast Sys C1000 chassis and 48 48 Dual Fast reac 185 1048 tem tion module system C1000 384 well System C1000 chassis and 384 well reaction mod 185 1384 ule system S1000 96 well Fast Sys S1000 chassis and 96 Fast reaction mod 185 2096 tem ule system S1000 48 48 Fast System S1000 chassis and 48 48 Dual Fast reac 185 2048 tion module system S1000 384 well System S1000 chassis and 384 well reaction mod 185 2384 ule system 5 O Real time detection modules CFX96 Real Time Detec Real time detection module with 96 well 184 5096 tion module block Software C1000 Manager soft Software package to run PCR with one or 184 4000 ware more thermal cyclers in a single window CFX Manager software Software package to run real time PCR 184 5000 and PCR with one or more thermal cyclers in a single window Accessories for the 1000 series Table 23 Catalog numbers for the 1000 series instruments and software number CFX Manager Security Software package to run real time PCR 184 5001 Edition software and PCR with one or more thermal cyclers in a single window Includes security fea tures for CFR part 11 compliance Accessories for the 1000 series 111 1000 Thermal Cycler Manual Appendix C Catalog Numbers The Table 24 lists accessory parts for the 1000 series instruments It in
112. step 4 is selected to delete Press ENTER to continue to the next screen In this example step 4 is selected to be deleted Choose DEL T T ee ees Hi Ai leit Ge 5 Step 4 3 65 0 H Selected 55 aly H r 5 a5 lt aan EDIT OFTION Delete the selected step Press ENTER to delete the step and continue to the next screen Notice that the deleted step parameters are replaced with the parameters of the next step In this example step 4 was deleted so step 5 parameters moved to step 4 and step 6 parameters moved to step 5 Step 5 5 aii parameters MEE cme S moved to Ez s i ia j a FOR Cz I es Confirm the deletion When prompted with Save Changes press the right and left arrow keys to select Yes or No Select Yes and press ENTER to delete the step Select NO and press ENTER to return to the beginning of this step 37 1000 Thermal Cycler Manual Create and Edit Protocols In this example YES is selected JL WL New step 4 Pt m iT P dn in iN piv oe ae a I m oT R N i H o i a P PT gm A bo ri 1 Select Yes or NO ee i 9 Edit a protocol step optional Choose EDIT to change the parameters in the existing protocol steps TIP When you first select EDIT you can edit the parameters in the selected protocol step To edit the parameters in a different protocol step press the s a i 2 HE HASSE CT TEn Change the first parameter
113. t use the thermal cycler will require some cleaning and upkeep Clean the base page 91 Clean the reaction module page 92 Replace the fuses page 94 TIP For robotic installations where many instruments run constantly use a regular program checking for dust spills and debris that could interfere with optimal instrument performance Cleaning the 1000 thermal cycler Base The 1000 thermal cycler should be cleaned on a regular schedule to remove any debris or dirt that might interfere with proper function Clean the base to prevent damage to the air intake or reaction module bay In general always use a soft cloth and water to wipe off spilled solutions and debris immediately NOTE For instructions on handling and cleaning radioactive or biohazardous materials consult the guidelines for radiation safety and biosafety provided by your institution These guidelines include cleaning monitoring and disposal methods for hazardous materials To clean the base follow these instructions paying careful attention to the warnings WARNING Prevent electrical shock Always turn off and unplug the instrument before cleaning it 91 1000 Thermal Cycler Manual Vaintenance amp Troubleshooting 92 e Clean the air vents Remove dust with a soft brush damp cloth or vacuum cleaner Remove any heavy dust that is deep in the vents with a vacuum cleaner Cleaning the vents allows sufficient air flow for precise thermal control during
114. the next target temperature Shorten the run time by minimizing the difference between target temperatures in successive steps To minimize target temperature differences add a temperature gradient Step 6 on page 23 Begin by optimizing the difference between the annealing and extension temperatures Use the results of this gradient experiment to determine the highest possible annealing temperature that does not sacrifice the PCR yield See Optimizing Temperature Steps With a Gradient page 55 for an example Finally choose an annealing temperature with the smallest temperature difference from the extension temperature Minimize the final extension step The final extension step completes the synthesis of amplicons Optimize this step when you need a high percentage of full length amplicons at the end of the PCR During each cycle the extension step is typically 30 seconds If amplification for 30 seconds is sufficient during cycling then a longer final extension step is unnecessary Optimizing Temperature Steps With a Gradient To minimize the final extension step Step 9 on page 26 choose a hold time between 30 seconds and 2 minutes for targets between 100 and 1 000 bp Then test the hold time you choose for sufficient amplification Alternatively add an increment page 28 to the extension step to test for the best hold time e Minimize the number of repeats in a GOTO step Minimize the repeats in the GOTO step to minimize th
115. this example the copy will be put into the GRANT folder STO Select a folder TIP If the folder you select is secure enter the password to copy a file into that folder 4 Enter a name for the copied file Type a new name for the protocol copied file Press the up and down arrows to select letters and press the number keys to type numbers Then press ENTER to accept the selection NOTE A protocol file name can contain from one to eight characters The characters are numbers or capital letters Each protocol name must be unique In this example the name of the copied protocol is STD2GR COPY STD TO Enter anew name Hew Hame ETOZGRE TIP After copying the protocol file make changes to the file by selecting EDIT in the main menu page 34 Moving a File With the MOVE function To move a protocol file to another folder select FILES gt MOVE Moving a file is useful when you want to move protocol files stored in the main menu to a folder you created TIP To secure a protocol file that was saved in the MAIN folder move the file to a folder that you created page 72 and secured page 71 To move a protocol file follow these instructions 67 1000 Thermal Cycler Manual Protocol Files and Folders 1 Select MOVE To select MOVE press the arrow keys Then press ENTER to confirm the selection and continue to the next screen FOLDERS PROTOCOLS 0P Select MOVE URE DELETE RENAME RENAME 2
116. tocol by optimizing the annealing temperature step by using a gradient page 55 Lower the lid temperature to reduce of sample loss due to evaporation The heated lid prevents condensation from forming in the microplate or tube which is critical when the sample volume is small However using the same lid temperature with a smaller sample volume can increase evaporation when the lid heat is the same To prevent evaporation lower the lid temperature and test the reaction for evaporation Transferring Protocols From Another Thermal Cycler Run the reaction in a 384 well reaction module The 384 well block is optimized for small sample volume and therefore this is the best block for this application Transferring Protocols From Another Thermal Cycler To transfer a PCR protocol from another thermal cycler to the S1000 thermal cycler with the same results you might need to lower the ramp rate If the results of identical reactions run on a thermal cycler with a slower ramp rate do not result in the same data then change the ramp rate Follow these suggestions when transferring a protocol Match the ramp rate of the other thermal cycler by changing the rate in each relevant step page 31 In general the ramp rate for the annealing step could be lowered when you move a protocol from a thermal cycler with a slower ramp rate to the S1000 thermal cycler Increase the amount of Mg in the reagents to help the primers anneal when the cond
117. ue to the next screen NOTE Enter an extension time between 60 and 60 seconds cycle in whole numbers and within the time limits of a temperature step from 1 second to 18 hours In this example the increment is 1 second The hold time for step 3 will increase by 1 second each cycle Enter the extend parameter 30 Adding INC EXT RATE and BEEP Parameters 4 Confirm the step parameters Press ENTER to confirm the increment temperature and finish entering the step parameters Press the right arrow key to select No and change the parameters for this protocol step Press ENTER to confirm the selection Changing the Ramp Rate RATE in a Temperature Step The RATE parameter changes the ramp rate of a temperature step The ramp rate is the rate at which the thermal cycler heats or cools to the target temperature of a step To change the rate follow these instructions 1 Choose OPTION after entering the initial temperature step parameters Press the right and left arrow keys to choose OPTION Then press ENTER to continue to the next screen In this example the OPTION will be added to step 3 parameters q5 4 AG Step 3 JS S aH selected BS n sA Choose OPTION Ho gies 2 Choose RATE to change the ramp rate Press the right and left arrow keys to choose the RATE parameter Then press ENTER to continue to the next screen FIR FOR FUR mr ie ee me ey Choose RATE IHC EXT 3 Enter a ramp
118. ults from the first wider temperature gradient Bracket the annealing temperature that you choose in Step 3 by 5 C Choose the final annealing temperature from the highest successful temperature in the results for this gradient For example if the highest successful annealing temperature in Step 3 is 60 C then bracket that temperature with a narrower gradient 5 C In this example the narrow gradient is from 55 to 65 C Choose the final annealing temperature from a successful high temperature in the gradient experiment as described in Step 3 on page 55 Optimizing Runs With Small Sample Volumes Running a protocol with a small volume requires optimization to prevent evaporation and condensation Follow these suggestions for optimizing PCR with a sample volume below 10 uL Read the following list for instructions about optimizing reactions with small sample volume 56 Control evaporation with a wax seal A wax such as Chill out Liquid Wax is the best seal for reducing evaporation Further reduction is possible by adding a second seal such as a cap or film in addition to the wax Reoptimize the annealing temperature to prevent nonspecific priming and increase target amplification When running an established protocol with a smaller sample volume lt 10 uL you might need to reoptimize the annealing temperature If you observe mispriming or low amplification that is not a result of reagent problems then adjust the pro
119. unctions Protocol file Folder rm SECURE functions functions HELETE RENAME REHAME Notice that there are two lists of options Use the options listed below FOLDERS to manage folders page 72 Use the options listed below PROTOCOLS to manage protocol files page 65 Table 13 describes all functions in the FOLDERS and PROTOCOLS lists Table 13 List of functions in the FILES option Function Description PROTOCOLS Copy an existing protocol file and save it with a new name For EA instructions see page 66 Move an existing protocol file to another folder For instructions MOVE see page 67 DELETE Delete an existing protocol file For instructions see page 68 RENAME Rename an existing protocol file For instructions see page 69 FOLDERS NEW Create a new folder For instructions see page 72 Protected files can be deleted and edited but only after the SECURE password protecting them is used For instructions see page 71 DELETE Delete an empty folder For instructions see page 73 NOTE A folder cannot be deleted if it contains protocol files RENAME Rename an existing folder For instructions see page 74 Copying Moving Deleting and Renaming Protocol Files The FILES option lists all the functions available for managing protocol files This section provides step by step instructions for using those functions e Select COPY to copy a protocol file next section page 66 e Select MOVE to move a
120. ure to store it in a safe location WARNING Damaging the ventilation fins on the bottom of the reaction module could compromise the ability of the module to heat and cool correctly 11 1000 Thermal Cycler Manual Get Started Opening and Closing the Adjustable Lid The inner sealing plate of the lid applies both force and heat which help run consistent and successful reactions for these reasons e Force on the reaction caps or sealing tape helps prevent evaporation e Heat on the caps or sealing tape prevent condensation For recommended lid heat and force parameters see page 33 The following two sections describe how to open and close the lid e Opening the lid below e Closing and adjusting the lid page 13 WARNING After a run the heated lid may be hot Use caution when opening and closing the lid Be sure to avoid touching the heated lid or block unless it has cooled Opening the Lid To open the lid follow these instructions 1 Release the lid force knob Twist the lid force knob in a counterclockwise direction to release the lid Lid force knob Turn knob to release seal and force 2 Lift the lever The lid lock is a lip on the lid To release the lid push back on the lid while lifting the lever Lid lever 1 lock unlocked prs 3 Open lid completely Continue lifting the lid lever toward the back of the reaction module so that it stays open independently 12 Control Panel Keys and Main

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