Team:Todai-Tokyo/Protocols/Colony PCR

Colony PCR Protocol

 * 1) Aliquot into PCR tubes 5µl of MilliQ
 * 2) Pick single colonies with a toothpick and immerse in the above MilliQ to suspend
 * 3) Put in PCR machine set at 95ºC for 5 min.
 * 4) Add to each tube 5µl of the following solution and mix by pipetting
 * 5) 0.1µl　100µM　5’ primer
 * 6) 0.1µl　100µM　3’primer
 * 7) 0.8µl　2.5mM dNTP
 * 8) 1µl　10x　standard buffer
 * 9) 2.92µl　MilliQ
 * 10) 0.08µl　Ex-Taq
 * 11) PCR using the following program
 * 12) 95ºC 2 min
 * 13) 95ºC 30 sec
 * 14) 52-55ºC 30 sec （Depending on the Tm of the primers）
 * 15) 72.5ºC 15 sec × （# of kb of DNA to be amplified）
 * 16) 95ºC 30 sec Repeat 2-4 29 times
 * 17) 25ºC pause