sequencing PCR products

Joseph Leo Wiemels wiemelsj at uclink2.berkeley.edu
Thu Jan 23 02:04:44 EST 1997


Amersham makes a PCR product sequencing kit in which two enzymes are used
to digest away the single-stranded PCR primers, then you can add your
sequenciing primer and do the regular sequenase reaction (except that
annealing of your primer is done by heating to 100 degrees C, then
quenching on ice).  It works great for me even when I use either of the
original PCR primers as a sequencing primer.  If you already have your
sequencing kit you can buy the two enzymes separately (check catalog)

Joe Wiemels
wiemelsj at uclink2.berkeley.edu
 
Sharon Lefebvre (slefebvr at umabnet.ab.umd.edu) wrote:
: Is it possible to directly sequence PCR products? To summerize the problem
: I'm trying to solve: I want to sequence a portion of both alleles of a
: gene. I've amplified it with no problems. I could clone the product and
: sequence, but then I'd be dealing with a clone, so if the original PCR
: product contained a mixture of two sequences (i.e. from two alleles), then
: I'd only be sequencing one at a time, so I'd have to look at many clones
: to find both alleles. Any ideas or suggestions?

: Sharon




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