Accuracy of PCR product seq.

Dr. Duncan Clark Duncan at nospam.demon.co.uk
Sun Mar 28 10:04:22 EST 1999


In article <7delfn$141 at net.bio.net>, radomski <radomski at unixg.ubc.ca>
writes
>Has anyone come across an article that deals with the overall accuracy of 
>sequencing PCR products? I just replied that the error is 
>probably the same as the error rate of the Taq used in the PCR.

Although the enzyme will have the same error rate as standard Taq it is
very doubtful that you will ever see the errors in sequencing unless the
error occurs in the first cycle or so of the linear amplification and is
then copied in the next 30 cycles. As you are amplifying linearly i.e.
30x that error will have had to be made on every amplified copy of the
template for it to be visible.  There will definitely be mistakes but
because you are not separating individual amplified molecules from one
another but viewing them all at once the odd error in an individual
molecule will never be above your background noise level. The signal to
noise ratio of correct to incorrect molecules is too high to see the
errors. 

Duncan
-- 
The problem with being on the cutting edge is that you occasionally get 
sliced from time to time....

Duncan Clark
DNAmp Ltd.
Tel: +44(0)1252376288
FAX: +44(0)8701640382
http://www.dnamp.com
http://www.genesys.demon.co.uk




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