PCR Amplification of DNA: Difference between revisions
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*Run PCR Program. Normally use touchdown PCR ('''DAVETD''') as follows: | *Run PCR Program. Normally use touchdown PCR ('''DAVETD''') as follows: | ||
#1 min at 94 | :#1 min at 94 | ||
#30s at 65 | :#30s at 65 | ||
#2 min/kb at 72 | :#2 min/kb at 72 | ||
#30s at 94 | :#30s at 94 | ||
#30s at 63 then -0.5/cycle | :#30s at 63 then -0.5/cycle | ||
#2 min/kb at 72 | :#2 min/kb at 72 | ||
#Repeat steps 4-6 28 times | :#Repeat steps 4-6 28 times | ||
#30s at 94 | :#30s at 94 | ||
#30s at 45 | :#30s at 45 | ||
#11 min at 72 | :#11 min at 72 | ||
# | :#Hold at 4 until ready | ||
*Purify PCR product if necessary using Qiagen kit (Add 5x PB) | *Purify PCR product if necessary using Qiagen kit (Add 5x PB) | ||