Chlamydia PCR cycling conditions - (Mar/09/2010 )
I'm fresh in running PCR. I have an enquiry about the PCR cycling program on detection genus of Chlamydia.
I'm using the primers stated its Tm is 56-58C.
however, the PCR cycling conditions are as follows
1. 10min at 94C
2. 15sec at 95C for 40cycles
3. 15sec at 55C
4. 30sec at 72C
I have no product for this conditions at the end. I'm thinking to do a gradient to measure the optimum temp for annealing
Please advice for other options. Thank you very much.
How big is your expected product?
Have you tried various dilutions of your template?
Do your PCR reagents (eg enzyme, dNTPs etc) all work with other reactions?