PCR mistakes

Clemens Suter-Crazzolara, PhD un691cs at genius.embnet.dkfz-heidelberg.de
Wed Jun 3 08:04:19 EST 1998


On Wed, 03 Jun 1998 12:20:45 +0100, Richard P. Grant said:

>It's a bit worrying that you're getting errors using a proof-reading
>enzyme.  I would suggest trying different enzymes.  

Yes, that was one of my ideas too. Perhaps I should try Pfu like you did.

>Silly question - how do you know you have the mistakes?  Are you comparing
>with a published sequence?

You address another good point. The sequence was published long ago, I have amplified 
it without apparent mistakes in the past, but now all of a sudden I find
PCR errors. So, I thought it were maybe sequencing errors, but the ABI profiles
look pretty convincing, and the mistakes appear to be distibuted
more or less throughout. I still haven't figured this out completely.


clemens




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