1. Is it normal to see only one distinct peak in the melting curve, but actually have several Tm values in the melt curve report? An example is shown below which I just copied and pasted from the machine's report:
Well Well Identifier Peak ID Melt Temp Beg. Temp End Temp
A1 sample1 A1.1 81.5 76.0 86
A1.2 74.0 69.5 75.5
A1.3 67.0 65.5 69
A1.4 62.0 59.5 64
2. I am also getting slightly different Tm values for the same PCR run, same species of the organism but different DNA samples. Example of the different Tms: 81.5, 81, 82.....Sometimes, I also get a Tm value of 81.5 for my no template control but the peak is only a slight bump on the melting curve with a Ct of >35. Does this mean I contaminated my negative control with my DNA?
Please help me iron out this mess.
Edited by pop09, 06 November 2009 - 08:27 PM.