Colony PCR

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℃ and then spin the condensation down in a microfuge. Set up the PCR reaction as follows:

5ul H2O + cells

5ul 5uM primer2

5ul 10×Taq Buffer

5ul 2mM dNTPs

0.5ul 10 mg/ml acetylated BSA

1ul Taq DNA polymerase

23.5 ul H₂O

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