Alm:PCR

Standard PCR reactions

• ~100 ng template
• ~100 pmole each primer
• buffer to 1X
• polymerase
• denature 94-95°C
• anneal 5°C less than lowest primer hyb temp
• extend 1’/kb to be amplified

PCR Master Mix (2.5X)

• 62.5 U/ml Taq DNA Polymerase
• 125 mM KCl
• 75 mM Tris-HCl, pH 8.3
• 3.75 mM Mg(OAc)2
• 500 uM each dNTP

5'-CATTAG can be useful to add to prevent exonuclease activity from degrading recognition sites