Question

How will you calculate annealing temperature for a specific gene of intersesr?

Answer 1

have been using gene specific primers for PCR. The primer synthesis report gave a lower Tm (by 7 degrees Celsius) than has been used elsewhere for the same primers. I have tried to optimize, but I continue to get 2 consistent bands; one of the expected size (about 800bp) and another smaller size (about 300bp). How can I solve this problem, so that I come up one clear band