TEGlab LAB USER MANUAL
Contents
1. pipette stock solution inside the fume hood Cast the gel and rinse off the tube or bottle Discard your gloves when you leave the electrophoresis laboratory After the gel run DO NOT leave power packs on and DISCONNECT cables from the electrophoresis apparatus When using the BioRad gel imaging system or the UV tables make sure your gel is dry and that you clean the light tables well Don t use wet gloves when typing Washing room 7109 A dishwasher an autoclave and a backup clean water machine are in this room Johanna takes care of operations here She collects the dishes from the laboratories If you have something special to wash or to autoclave talk with her Teaching laboratory 7119 Ethanol storage 2 scales and a pH meter are here ADDITIONAL NOTES Ethanol Christina Nokkala Ethanol is stored in an assigned locked cupboard in teaching laboratory 7119 Keys are in Christiina Nokkala s possession Ask her for ethanol or Ville if she is not around Fridges freezers and the cold room The lab has plenty of freezer space due to the fact we have our own freezer room 7111 Defrosting of standing freezers will be organized at least once a year Ville will inform you about this on the email list Dishwashing Johanna room 7109 Used lab ware should be rinsed and taken into the basins placed near the sinks in every laboratory Johanna collects them and does the actual dishwashing If you have something special to wash or a2. Blocker Casein trypsic peptone tryptone Calcium Chloride anhydrous Calcium Chlorid Dihydrat Citric acid monohydrate sitrunahappo 1 hydraatti Certistain CTAB Cetyltrimethyl ammonium bromide Chloroform Chromosulfuric acid Carmin fo microscopy Colchicine Canada Balsam in Xylol Neutral trans 1 2 Diamino cyclohexane N N N N tetraacetic acid Dithio oxamide rubeanic acid Deoxycholic acid Sodium Salt 3d 12d Dixydroxy 5b Cholan 24 oic acid N N Dimethyl Formamide Dimethyl Sulfoxide EDTA Dinatriumsuola TITRIPLEX Entellan Ethylene Glycol bis b aminoethylether N N N N tetraacetic acid Ethylenediaminetetraacetic acid Ethylene Glycol TriEthanolaminhydrochlorid Ether Diethyl Ether Eetteri Etan 1 2 Dion 4096 i vatten Etaani 1 2 Dioni 40 vesiliuos Glyoxal 40 wt solution in water Eosin gelblich Exsikkatorfett Formamide Formamide Ultra pure grade Formic Acid 9896 Formaldehyde solution Fuchsin Basic D Gilucose Dextrose Corn sugar D Glucose anhydrous Glycine Gelatine Glycerin etwa 87 Glycerol L Glutamic acid D Galactose Hepes Hydrochloric acid 36 38 Suolahappo Hydrochloric acid 2 0 N 2 0 M Hydrochloric acid 0 1 N 0 1 M Jod doppelt sublimiert Kaliumchlorid Potassium chloride Kaliumdihydrogenphosphat Potassium Dihydrogen Phosphate Kaliumcarbonat Potassium carbonate di Kaliumhydrogenphosphat Trihydrat Kaliumdisulfit Potassium disulfite Kalium Chromi I
3. This room also contains the lontorrent OT2 which are next generation sequencing machinery Meri Lindqvist will tell you more about the possibilities of using it This room also contains 4 deep freezers 1 is an emergency freezer Ask Ville for space Sequencing and Fragment Analysis Tatjana Saarinen is in charge of preparing adding formamide amp size standard in fragment analysis and running sequencing and fragment analysis samples Preparation and handling of EtBr agarose gels TEGlab encourages people to use safer gel dyes like Midori Green which is ours to use in the gel lab fridge But if you insist on Et Br The ethidium bromide EtBr used for staining of agarose gels is mutagenic it can directly incorporate into DNA and change its structure EtBr is harmful if inhaled swallowed or in skin contact It is soluble in water and thus can be removed from the skin by rinsing with water and mild detergent When using EtBr in the lab you must 1 prevent its evaporation 2 prevent contact with your skin by always using gloves when handling items possibly contaminated with EtBr 3 protect others by not spreading EtBr anywhere outside the apparatus directly involved in the gel run Always dissolve the agarose in the microwave first and let the agarose cool off in a Erlenmeyer bottle under running water or using magnetic stirrer before adding one drop of EtBr 2 5ul 50ml gel solution stock solution s concentration is 10mg ml
4. wear gloves when you are working here and dispose them when you leave the room TEGlab encourages people to use safer dyes such as Midori Green provided in the fridge With the UV tables use BOTH goggles and protection mask if you are working in direct UV exposure If you want cut bands from your gel use the Dark Reader Don t cut gels on the Gel Imaging system Cold rooms 7113 amp 7071 Shelf space in cold room is allocated according to the needs of the groups If you need storage place there contact Ville RNA laboratory 7066 Clean DNA and RNase free space for working with RNA samples DO NOT move any pipette sets from here or to here Also new pipette sets should be autoclaved before placing here Erica is in charge of this area A TissueLyser for preparing samples is also here ask PnP research group for permission to use ABI Room 7075 Keep this laboratory s door closed because of air conditioning Do not run the sequencer during thunder storms the laser might get damaged The lab has ABI Prism 3130xl Genetic Analyzer 16 capillary sequencing machine You can leave your ready sequencing samples to Tatjana or once you have received proper training operate the machine yourself NOTE Use ONLY SEMI SKIRTED 96 Detection Plates in this machine otherwise there is risk of capillary damage This room also contains Thermo centrifuge which is able to spin truly fast rpm at low temperature Consult Mikko Nieminen
5. 1 Propanol 2 Propanol Isopropyylialkoholi 1 4 Bis acryloyl piperazine for electrophoresis PVP K30 Polyvinylpyrrolidone PVPP Polyvinylpyrrolidone Polyethylene Gilycol Penicillin streptomycin solution Quinacrine Dihydrochloride SDS Sodium dodecyl sulfate Saccharose 5 Sulfosalisyylihappo IN CASE OF AN EMERGENCY D Sorbitol anhydrous Silvernitrat PA Starch Potato Silica gel 60 0 015 0 040 mm Silica gel with moisture indicator blue gel for drying TRIS hydroksimetyyli aminometaani Triton X 100 Iso octyl phenoxy polyethoxyethanol a Terpineol Trypsin 1 250 Titriplex Il Tween 20 UREA Xylene Yeast extract Common emergency telephone number 0 112 Police Pharmacity HelpDesk 0 100 22 0 0800 157 777 Fire extinguishers are located along the corridors Emergency shower is on the corridor next to Pre PCR Get acquainted where it is before starting to work in laboratory FIRST AID Kits can be found from almost all the labs Chemical Safety Instructions are found from the teaching lab chemical storage and from the network drives ChemDAT database Path utu i verkkolevyt Biologian genetiikka Kemikaalilista Chemical Data Sheets In case of 20 freezer breakdown 1 Check the temperature If the alarm is beeping the temperature is above 14c If it keeps on rising start to move the items to other freezers 2 Use the freezer room 7111 to evacuate Notify Ville an
6. 4597 30 updated 16 1 2014 EMERGENCY INSTRUCTIONS ON THE LAST PAGE TEGlab members and phone numbers The area code for all phone numbers is 02 Key personnel Ville Aukee Laboratory manager 333 7083 Tatjana Saarinen Laboratory master 333 5548 Raija Rouhiainen Secretary 333 5575 For additions and or corrections to the manual contact ville aukee Qutu fi EMERGENCY INSTRUCTIONS ON THE LAST PAGE LAB RESPONSIBILITIES COMMON CHEMICALS ORDERS TATJANA SEQUENCING ABI MAINTENANCE ORDERS TATJANA ETHANOL CHRISTIINA DISHWASHING AUTOCLAVING JOHANNA GMO WASTE NOTIFY JOHANNA HAZARZOUS WASTE TATJANA BALANCE AND PH AREA TATJANA PRE PCR SHARED CLONING AREA USER CLEANS RNA ROOM ERICA GEL ROOM TATJANA ION TORRENT PREP MERI Responsibilitites include supervising the tidiness of the assigned area reporting of problems or malfunctions to TEGlab manager advising in use of equipment and space etc However lab users are always expected to clean after themselves EMERGENCY INSTRUCTIONS ON THE LAST PAGE First things to do when starting in the lab e Read this manual e Introduce yourself to Ville room 7011 e Join the TEGLab mailing list Go to https Aists utu fi mailman listinfo teglab and fill in the form e Fillin the TEGlab user license form ask Ville Some rules of the lab e Attend the regular TEGlab meetings about every first Wednesday of each month and learn about new things or discuss issues You must have
7. F CHEMICALS FOUND IN THE LAB If you use see that any chemical is running low please inform Tatjana Saarinen It can take up to a month for certain chemicals to arrive so let her know well in advance All the chemicals are in the teaching laboratory in a locked room If you re looking for something special best to ask Tatjana first room 7058 Phenol chloroform isopropanol etc are stored in the fume hoods or in a fume cabinet in teaching lab Updated list of the common chemicals is in utu i verkkolevyt Biologian genetiikka Kemikaalilista Detailed information about each chemical s toxicity safety and handling information etc can be seen in ChemDAT database also in our network drive Path utu i verkkolevyt Biologian genetiikka Kemikaalilista Chemical Data Sheets There are also the safety notions in paper form in the pre PCR lab Unfortunately those are in Finnish only More information about the safety notes of all chemicals can be found from ICSCs pages Agar bacteriological Agar Corn Meal Agarose low EEO iso Amyylialkoholi Ammonium Sulfate Ammonium persulphate Ammoniumeisen iii Sulfat Ferriammoniumsulfat 3 Aminopropyl triaethoxysilan 98 Ampicillin Sodium Salt Aktivkohle Alcian Blue 8 GX Acridine orange Azure B certified Acrylamide Bis Acrylamide Albumin Acetone Acetic Acid 99 100 Etikkahappo Boric Acid Boorihappo Bariumhydroxid Octahydrat Bromophenol Blue Blotting Grade
8. II Sulfat Potassium Chromium llIl Sulfate Kaliumnitrat Kaliumacetat Potassium Acetate Kaliumhydroxid Kaliumjodid neutral Kuparisulfaatti Koboltti II kloridi heksahydraatti Koboltti II Nitraatti Kamferia Ksyleenisyanoli Xylene cyanol F F DL Lactic acid sodium salt Lactic acid about 9096 Maitohappo Litiumkloridi Lithium chloride Methyl 4 hydroxybenzoat Nipagin M Magnesiumsulfat Heptahydrat Magnesiumacetat Tetrahydrat Magnesiumchlorid L Malic acid Maleic acid Maleiinihappo D Mannose Mangan II chlorid tetrahydrat Methyl green Methyl a d Mannopyranoside N N N N tetramethylethylen diamin Temed Methylene blue trihydrate Metyleenisini MOPS 3 N Morpholino propane sulfonic acid Milk Non Fat Dry Milk Mowiol Methanol Natriumhydroxid Sodium Hydroxide tri Natriumcitrat Dihydrat Natriumfosfaatti Sodium phosphate anhydrous Natriumhydrogenphosphat Sodium Hydrogen Phosphate Anhydrous di Natriumhydrogen phosphat Dihydrat Natriumchlorid Natriumhydrogen carbonat Natrium carbonat anhydrous Natriumdihydrogen phosphat Monohydrat Natriumacetat Trihydrat Natriumsulfit Natriumiodid Natriumasetaattianhydraatti Sodium acetate anhydrous di Natriumtetraborat Decahydrat Natriumnitrat Natriumthiosulfat Pentahydrat Natriumborhydrid Natriumcacodylat Naphthol Green B Orcein Phenol Phenol Saturated solution Paraformaldehyd Pepton aus Casein n Propyl Gallate anhydrous
9. d then everyone by mailing teglab lists utu fi about the incident In case of 74c freezer alarm Call these people until someone answers and also read the instructions how to proceed VILLE AUKEE 050 3204118 SAIJA KILJUNEN 0445382408 MIKKO NIEMINEN 050 5671636 ERICA LEDER 050 3398204 ANTI VASEMAGI 045 3585410 HEIDI VIITANIEMI 040 7045585 HANNU M KINEN 0400 691338 SIIM KAHAR 046 5559427 MATTHIEU BRUNEAUX 044 9197667 MERI LINDQVIST 050 5850694 Instructions for an emergency situation 75 C Freezer Alarm goes on at 65 oC normal running temperature is 75 oC The alarm is connected to a remote system and people in a security company will see the alarms They will also start to call people on the list above Check the alarm codes from the display see instruction manual Check that the door is properly closed Do not open it if not then close it and wait for a while to see if the temperature returns to 74 C If yes then you re good to go home Remember the freezer temperature doesn t change quickly so monitor it for quite a while 15 min and see which direction it is going if the door is properly closed but the temperature keeps on increasing follow this procedure Move all the items from the broken freezer into the EMERGENCY FREEZER which is the 2nd freezer from the door It should be empty maybe some containers with dry ice so all the items will fit The mergency freezer has been set to 70 so decrease
10. filled out the lab user license before starting Be sure you have an assigned bench where to work Subscribe to TEGlab mailing list see above for instructions Keep you bench place clean and be tidy elsewhere too Be safe A laboratory can be a dangerous place Inform Ville at least 2 working days before vacating your bench o clear out your bench amp freezer storage COMMON SPACE AND EQUIPMENT If needed ask Ville for specific instruction manuals for each piece of equipment Certain people will also be able to help you with operation of certain equipment refer to the lab user license If something breaks down or is otherwise out of order contact Ville When ABI is in question you may also contact Tatjana Pre PCR lab 7114 Each bench section should have one vortex and minicentrifuge DO NOT move any machines without consulting Ville first Please DO NOT shift items or equipment between pre and post PCR sections Ask Ville about possible freezer space in this section of the lab DNA extraction hood in 7114 There s a vacuum pump and a manifold for DNA extraction in this hood Consult Katja or Kristiina in the use of these Ultrapure water The main ultrapure water system is in machine room 7114 pre PCR The backup clean water machine is in dish washing room 7109 RO Water See instructions for the simplest way to operate the water machine How to use PureLab Utlra machine Manual Dispense Step 1 Product Quality When yo
11. pendorf tubes used with these chemicals should be in a separate bottle If there is no bottle for that chemical you are using ask suitable bottles from Johanna Any waste containing bacteria should be placed to a RISKIPAK box yellow plastic bag It is essential that you clean up after yourself after using the fume hoods LIST OF COMMON EQUIPMENT Sequencing e ABI Prism 3130x genetic sequencer 16 capillaries analysis software GeneMapper 4 0 e lonTorrent OneTouch PCR 2 x Epperdorf Mastercycler Gradient 96 well 1 x MJ Research PTC 100 96 well 3 x Applied Biosystems 2720 Thermal Cycler 96 well 1 x Finnzymes Piko Electrophoresis e lots of small and large apparatuses Gel documentation pictures manipulation e BioRad GelDoc System Cloning e fully equipped cloning laboratory 7069 Talk to Ville First Cell culture e incubators fully equipped cell culture laboratory Spectrofotometry e NanoDrop ND 100 e Agilent 8453 Dell prosessor DNA extraction e 2x Eppendorf Vacuum Manifold salt extraction Cold storage e HERA Deep Freeze 85 C 3 x deep freezers 20 C freezers 20 freezer room refrigerators 4 cold room x 2 in addition e laminars fume hoods large and small centrifuges heat blocks ovens analysis and cup scales ice machine UV tables DarkReader incubators autoclaves PureLab Ultra Water system microscopes light and preparation RNA lab TissueLysery etc LIST O
12. the running temperature to 75 In case of a GMO related incident In a situation of unexpected contamination exposure or other serious incident occurs the following procedures will be followed 1 Prevent additional danger according to possibilities In a serious case contact the authorities immediately tel 112 Perform CPR if someone is hurt lsolate the danger zone inform the people working in the laboratory genlab lists utu fi teglab lists utu fi If necessary inform the surroundings Start to eliminate the danger decontamination of facility machinery protective clothing Repair the damages and find out what happened Find improvements safety procedures Responsible people in a case of GMO incident lab manager Ville Aukee room 701 1 tel 0503204118 laboratoriomestari Tatjana Saarinen room 7085 GMO facilities class 1 rooms 7069
13. u see only this L4 PRESS THE POWER BUTTON ONCE AND WAIT UNTIL THE SCREEN CHANGES ABOUT 30 SEC look for value 18 20 1 ENSURE PURELAB Ultra is in recirculation Step 2 Dispense 1 PRESS Dispense Controller once Cual 2 ROTATE Dispense Controller clockwise If the Dispense Controller is not rotated within 15seconds the unit will revert to recirculation mode 3 ADJUST the flow from the unit by rotating the Dispense Controller as necessary Note To prevent accidentally leaving the unit on continuous droplet dispense the PURELAB Ultra will only dispense for a maximum of 4 minutes from the last turn of the Dispense Controller For large volumes it is recommended that the Volumetric Dispense mode is used 4 PRESS Dispense Controller once to stop dispensing and return to recirculation mode Post PCR room 7116 This room contains the PCR machines and some bench place to prepare post PCR samples Note that the PCR machines have a reservation system Keep as few personal PCR programs as possible in the machines DELETE all programs which you are not using anymore The fancy looking BioRad PCR machines are not for common use lonTorrent related material is also located here the benches next to the PCR machines This room also contains the Electrophoresis laboratory Agarose electrophoresis apparatuses UV tables Dark Reader and the GelDoc imaging system are here SAFETY NOTES Some still use Ethidium bromide so always
14. utoclave discuss with her Keys to corridor Raija the secretary can provide you with keys Or if you already have university s electronic key notify number of your key to her to get access activated pH meter room 7119 Always consult Tatjana before using the pH meter for the first time Instruction sheets in Finnish and in English are next to the machine on the wall The pH meter is always turned on if it is turned off it has to be recalibrated Calibration solutions are next to the pH meter Pipettes Tatjana Saarinen room 7058 Tatjana Saarinen takes care of all common pipettes marked TEGLab She also calibrates and replaces broken parts to them If you have some problems or you are not sure of some pipette s calibration contact her NOTE there are also pipettes which are only for certain groups to use clearly marked Ask permission to take such a pipette Postal service In the top shelf of the mail pigeon holes you can find a locker for university internal mail sis inen posti and outgoing mail ulkoinen posti When expecting a special delivery such as 20 C or 4 C package give a forewarning to secretary and or laboratory master You may also make an announcement on the teglab mailing list Fume hoods and hazardous waste Hazardous waste is collected in bottles in the fume hoods Phenol chloroform iso amylalcohol and isopropanol are poured into the collecting bottles assigned to this and dirty pipette tips and ep
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