IGEM:MIT/2006/Notebook/2006-7-28

Amazing News
Transformants on every plate!!!

To do (am)

 * 1) Miniprep 5 liquid cultures, make glycerols, and send DNA for sequencing -complete
 * 2) PCR pchBA -complete

pchBA PCR

 * 1) Lets try to amplify out pchBA together and seperately
 * 2) 3 test tubes + 1 control
 * 3) Dilute new Invitrogen pchBA primers into working stocks of 25μM
 * 4) Tube 1 (pchBA):
 * 5) *49 &mu;L PCR supermix
 * 6) *1 &mu;L pME3366 template (1 nanogram)
 * 7) *.6 &mu;L pchB Forward (300 nanograms)
 * 8) *.6 &mu;L pchA Reverse (300 nanograms)
 * 9) Tube 2 (pchB):
 * 10) *49 &mu;L PCR supermix
 * 11) *1 &mu;L pME3366 template
 * 12) *.6 &mu;L pchB Forward
 * 13) *.6 &mu;L pchB Reverse
 * 14) Tube 3 (pchA):
 * 15) *49 &mu;L PCR supermix
 * 16) *1 &mu;L pME3366 template
 * 17) *.6 &mu;L pchA Forward
 * 18) *.6 &mu;L pchA Reverse
 * 19) 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)

 * 1) PCR cleanup pchBA PCR products and run 5 &mu;L on a gel to see if successful and put in -20 fridge
 * 2) make 3 liquid cultures (A,B,C) from each (8) transformant plate and put in 37 room