Taq slippage in PCR

Shiao Y. Wang sywang at whale.st.usm.edu
Sat Oct 8 23:35:53 EST 1994


John H McDonald (mcdonald at strauss.udel.edu) wrote:
: I'm trying to PCR and then directly sequence some DNA with a couple of
: runs of 7 to 10 T's.  I'm having problems, because "slippage" causes

I'm not quite sure what you mean. Are the runs of T's part of your
sequencing primer? Or are they in template DNA itself? I had a little
trouble sequencing cDNA because of the polyA tail. (I use cycle
sequencing with amplified DNA as template). I only had the problem when
the primer was immediately upstream of the polyA tail. The longer the
polyA tail, the worse the problem (one clone with 34 was almost
impossible, ones with less than 20 were okay). My solution was to get
readable sequence and then immediately make new primers just downstream of
the polyA region. Good luck.

Shiao Wang
University of Southern Mississippi




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