Wrong PCR product after Bisulfite Treatment-reason? - (Aug/24/2007 )
My Bisulfite PCR suffers from an error that I cannot explain and unfortunately cannot overcome.
I optimized my protocol for amplifying a particular region from genomic bisulfite converted DNA. It perfectly worked for months without any problems.
Suddenly I did no longer obtain a 300 bp band (as before) but a 200 bp band, although I haven't changed anything!! amplification of another region from the same template works.
I checked the primer sequences - they are ok. I ordered new primers, just in case the old ones were degraded - still obtain the wrong band.
I cannot think of any reason why my PCR out of a sudden doesn't work anymore. I am sequencing the "wrong" band at the moment, maybe this will give me any clues.
Can anyone think of a solution or has any suggestions how to overcome that problem? I would be very thankful
Thanks in advance,
Does your DNA come from the same samples? If you are using cells, maybe you have something like a repeat-insertion polymorphism? Okay, doesn't sound likely, but I have no other idea.