We are working on HCV PCR using degenerate primers. Our forward primer contains degeneracy at 4 positions. no degeneracy at 3' end. sometimes we get very bright bands and the very next day band intensity gets very low and even sometimes we don't get any band at all. our cDNA is good as we checked by real time PCR. Initially we were using DEPC water for PCR and DEPC treated PCR tubes for PCR using cDNA. now we used milipore autoclaved H2O , but no band. what could be the reason ??
problems in PCR with degenerate primers
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