Colony PCR

Colony PCR David Amberg This procedure will work for both yeast and E. coli:

Take a small colony and suspend it in 5ul of H2O in a PCR tube. Heat for 5 min at 95¡C and then spin the condensation down in a microfuge. Set up the PCR reaction as follows:

5ul H2O + cells
5ul 5uM primer2
5ul 10XTaq Buffer
5ul 2mM dNTPs
0.5ul 10 mg/ml acetylated BSA
1ul Taq DNA polymerase
23.5 ul H2O

PCR Conditions: 94¡C x 4min. then 35 cycles of: (94¡C x 1 min then 55¡C x 1min then 72¡C x 3min) followed by 72¡C x 20 min and a 4¡C soak. (5ul run out on a mini gel should be sufficient to see product.)