IGEM:MIT/2006/Notebook/2006-7-28: Difference between revisions
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''Transformants on every plate!!!'' | ''Transformants on every plate!!!'' | ||
==To do== | ==To do (am)== | ||
Miniprep 5 liquid cultures | #Miniprep 5 liquid cultures, make glycerols, and send DNA for sequencing -'''complete''' | ||
#PCR pchBA -'''complete''' | |||
==pchBA PCR== | ==pchBA PCR== | ||
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#*.6 μL pchA Reverse | #*.6 μL pchA Reverse | ||
#Run the tubes on the thermocycler at: 95deg for 3:00, then cycle through: (a) 94deg for :30 (b) 55deg for :30 (c)68deg for 2:15, then 72deg for 10:00 | #Run the tubes on the thermocycler at: 95deg for 3:00, then cycle through: (a) 94deg for :30 (b) 55deg for :30 (c)68deg for 2:15, then 72deg for 10:00 | ||
==To do (pm)== | |||
#PCR cleanup pchBA PCR products and '''run 5 μL on a gel''' to see if successful and put in -20 fridge | |||
#make 3 liquid cultures (A,B,C) from each (8) transformant plate and put in 37 room | |||
Latest revision as of 08:58, 28 July 2006
Amazing News
Transformants on every plate!!!
To do (am)
- Miniprep 5 liquid cultures, make glycerols, and send DNA for sequencing -complete
- PCR pchBA -complete
pchBA PCR
- Lets try to amplify out pchBA together and seperately
- 3 test tubes + 1 control
- Dilute new Invitrogen pchBA primers into working stocks of 25μM
- Tube 1 (pchBA):
- 49 μL PCR supermix
- 1 μL pME3366 template (1 nanogram)
- .6 μL pchB Forward (300 nanograms)
- .6 μL pchA Reverse (300 nanograms)
- Tube 2 (pchB):
- 49 μL PCR supermix
- 1 μL pME3366 template
- .6 μL pchB Forward
- .6 μL pchB Reverse
- Tube 3 (pchA):
- 49 μL PCR supermix
- 1 μL pME3366 template
- .6 μL pchA Forward
- .6 μL pchA Reverse
- Run the tubes on the thermocycler at: 95deg for 3:00, then cycle through: (a) 94deg for :30 (b) 55deg for :30 (c)68deg for 2:15, then 72deg for 10:00
To do (pm)
- PCR cleanup pchBA PCR products and run 5 μL on a gel to see if successful and put in -20 fridge
- make 3 liquid cultures (A,B,C) from each (8) transformant plate and put in 37 room
