50 µl reaction

• 5 µl 10x PCR buffer
• 2.5 µl 25 mM MgCl2
• 1.25 µl 10 mM each dNTPs (4 x 10 µl 100 mM dNTP (each) + 60 µl H2O)
• 1 µl 100 µM 5' oligo (i.e. 30 nmol dry in 300 µl H2O)
• 1 µl 100 µM 3' oligo
• 0.3 Taq polymerase (Amplitaq Gold for high fidelity)
• 39 µl H2O

Always prepare N+2 reactions (+ and - control)

Thermal cycles:

95 °C	95 °C	55 °C	72 °C	72 °C	20 °C
10 min	30 sec	40 sec	40 sec	5 min	indefinite
		25 x

This can be used to clone, mutate bases (see mutagenesis), check minipreps (touching colonies),
..... and anything else you can imagine.