About DNA sequencing

Tom Anderson via methods%40net.bio.net (by ucgatan from ucl.ac.uk)
Sun Oct 7 14:38:10 EST 2007


On Sun, 7 Oct 2007, kou wrote:

> After sequencing, entire sequence is not accurate. stretch of sequence
> immediately after primers is not accurate. So my question is How to
> choose accurate sequence ?

Your sequencing results should come with some kind of quality information
that tells you how much confidence you can put in each base. You can use
this to cut out the trustworthy stretch of sequence - the usual rule of
thumb is to take everything with confidence of 99% or over.

Also, any sequencing operation worth its salt will send you the original
chromatogram trace files, and with these, you can run your own
basecalling, which will give you both sequence and quality output. Getting
this set up can be a bit awkward, though: for the two best basecallers,
phred and lifetrace, you have to email their authors for source code, and
then compile it yourself. Well, or pay money for it. If you want to do
this, the thing to do is to find your local bioinformatics person and ask
them about it.

tom

-- 
Tom Anderson, MRC Laboratory for Molecular Cell Biology, UCL, London WC1E 6BT
(t) +44 (20) 76797264   (f) +44 (20) 76797805   (e) thomas.anderson from ucl.ac.uk


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