Standard PCR Reaction

| [10] primer | 10x dNTPs | 10x Buffer | 100x taq

Whole-Cell PCR

• Grow fresh overnight cultures of the cells to be used as template in LB.
  Important: Do NOT use E. coli grown in Davis Minimal (DM) media as template for whole-cell PCR. Ingredients in DM interfere with reliable amplification. (Probably the high concentration of citrate added as a chelating agent). Amplification from cultures frozen is also not as reliable as from freshly grown cells.
• Dilute cells 100-1000x fold into ddH₂O (NOT saline) in a 1.7 ml Eppendorf tube or 96-well microplate.
• Add the diluted cell mixture as 1/10th the final volume of the PCR reaction.
• Add an initial denaturing step of 10 minutes to your PCR program at 94°C to lyse the cells.