PCR sequencing problems

sequencing hbbio305 at email.csun.edu
Mon Oct 20 12:34:11 EST 1997


I have been sequencing short to medium lenght PCR products successfully
(on a 377 ABI) up until a few weeks ago.  I use shrimp alkaline
phosphatase and exonuclease I as a cleanup.

Recently all PCR sequencing has come to a halt.  I get only short blips
on the gel image.  As well, the controls have much higher background,
and more N's than normal.  

I have tried increasing the DNA and ethanol precipitation of the DNA to
remove the inactive enzymes, to no avail.

Has anyone else had this problem?

Are there any suggestions?

Jan




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