ikhan at BGSM.EDU
Mon Aug 26 16:03:06 EST 1996
I am sequencing PCR product by using sequenase version 2.0 DNA sequencing
kit (USB, Cleveland, OH). The PCR product is obtained by amplifying the
cDNA (from human T cells). In a particular segment of gene of interest
the findings are as follows- when I used forward primer for manual
sequencing, sequence is completely identical in healthy control and
patient samples and matches to the published cDNA sequences. On the
contrary, when I use reverse primer for seuqencing, two 'T's are missing
in patient sample while the healthy control sequence matches to published
sequence. The sequencing autoradiograms are very crisp and clear and I
can read about 200 bases with ease.
I am using the alkaline denaturation method and following the
I am just curious that these missing bases are just PCR artifact or
something else. Please advise me.
Islam Khan, Ph.D.
ikhan at bgsm.edu
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