Colony PCR

Transfer few cells from a colony to the bottom of a PCR tube. Add 100uL of sterilized water to the tube and microwave it at max effect for 2 mins to destroy the cells.Use 2uL of the colony DNA template suspension in the following PCR mix.

PCR Mix:

5μl Dream Taq Buffer
1μl dNTP
1μl Primer1 (VF2)
1μl Primer2 (VR)
3μl 25mM MgCl2
1μl Dream Taq Polymerase
2μl Template suspension
36μl sterile destilled H2O

PCR Program:

 
95°C for 2minutes
30 cycles:
95°C for 1 minut  
melting temp -5°C for 30seconds 
72°C for 1 minute/kb
72°C for 10 minute 

4°C on hold