PCR Using T7 and SP6 Primers

Andrey S. Shaw shaw at immunology.wustl.edu
Sat Aug 2 22:18:12 EST 1997


I have used these primers at 55 degrees annealing with no problem.  The
algorithms that are used to calculate melting temperatures are very
crude and not very applicable to real life situations.  in general, I
always use higher temperatures than the calculated Tm.  
	A better solution is to slightly extend the sequences of the T7 and Sp6
primers to 20 -22 bases.  That is what we have done and now I anneal at
65 degrees.

andrey shaw



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