Anyone familiar with real time PCR?

Question

We recently purchased a new primer/probe set from AB which uses the TaqMan kit. When I ran some samples to test it, the curves came out weird. They were almost perfect bell-shaped curves. I used the same sample but in different concentrations. The less sample I added , the wider the curves. I ran them in triplicates and each curve was almost exact. Any ideas on what could be the problem? thanks

Answer 1

Call ABI's tech support.  Biotech company support lines are extremely knowledgeable and helpful.  You'll get much better help from them than trying to communicate through email.

Make sure you run everything the way their protocol has it (at least for these test runs) from set up, loading, to software calibration (make sure you have it set up for TaqMan and not SYBRGreen).