Problem with PCR
Varada P. Rao Memorial Univ. of Newfoundland St. John's Canada
vprao at kean.ucs.mun.ca
Sat Dec 17 21:49:47 EST 1994
I have experienced of late a new problem in addition to the weak signals
that I often get in my PCR. I perform PCR to amplify T cell receptor cDNA
using several Vb-specific primers but using a single constant region primer.
A year ago I have done initially some PCRs to deterimine optimal conditions
for my work. After that, I recently started again to repeat the previous
observations. All the reagents and primeres were stored in -20 Celcius.
Here comes the problem:
I do get the appropriate sized bands specifically ( negative controls do
work) but now, with almost all the appropriate Vb-primers + the same
constant primer, I get " doublet-bands"; one of the correct size and
the second one is smaller by 50bp approx. This pattern is constant in every
This double-band problem is never seen with beta-actin primers under
I am really going mad with this. I would greatly appreciate any input or
Thanks very much in advance.
"vprao at kean.ucs.mun.ca"
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