What about the contamination in negative control with higher Tm than target gene - (Oct/05/2007 )
My nagative control comes out at about 30 cycles. There are two peaks in the melt curve: one is similar to the target gene, which I believe is contamination, the other peak comes out at about 2 minutes after Tm of target gene. Could any one help me figure out what that is ?
First I guess you mean two degrees higher (instead of two minutes)? Second, we would need to know what your negative control consists of? Is it the mastermix plus water instead of template? Are you doing PCR or RT-PCR? If it's and RT-PCR, and your negative is an RT-, it could be contamination with gDNA. Have you run a gel to check the length of the product?