Transcription, chromatin and DNA repair > Resources > Protocols > Colony PCR

Yeast colony PCR (up to 2 kbp).

1. Using the tip of a toothpick, transfer a small amount of cells to the bottom of a PCR tube.
2. Microwave for 1 min at max power together with a beaker of water (to absorb excess energy).
3. Place on ice.
4. Prepare reaction mix:   
   
   

   H2O	12.75 µl
   10xBuffer	2 µl
   20 mM MgSO4	1 µl
   2 mM dNTPs	2 µl
   Primers (20 µM)	2 µl (1 µl each)
   Taq (1 unit)	0.25 µl

   
   
   
   
   
   
   
   
   
5. Mix on ice and add to PCR tube.
   
6. PCR:
   
   

   96°C	2 min
   30 cycles of:
   94°C	1 min
   52°C	30 sec
   72°C	2 min
   4°C	forever