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==Materials==
*Primers – order from IDT prediluted to 100 mM in TE. Make working solution of 1 uM (100x) in water with both sense and antisense primers combined(300uL water, 3uL of each primer)*dNTPs – dilute to 2 mM each in water, make single use aliquots (50 uLaliquots... 10uL of each dNTP, 460uL water)
*Template – generally 1uL or less of a plasmid miniprep
*Polymerase – use Pfu Turbo for cloning and Taq for noncloning. Use appropriate buffer.
==Protocol==
*Use the following volumes per reaction
::*Buffer, 5 uL of 10X buffer (Dave's fridge)
::*Primers, 10uL of 1uM stock solution in water (both primers combined)
::*dNTPs, 5uL of 2 mM ("molecular biology stuff" box in freezer)
::*Sterile water, 28 uL
::*Template 1 uL
::*Polymerase 1 uL(turbo pfu found in "enzymes" box in freezer)
*Run PCR Program(approx 3.5 to 4 hours). Normally use touchdown PCR ('''DAVETD''') as follows:
:#1 min at 94
:#30s at 65