Hi

try designing new primer pairs. Or if available use already published ones. If its just for normalization purposes try to use another housekeeper.

Regards

I agree that it is probably your primer pair.  You could go further outside your region of interest with another primer pair that is more closely matched as far as Tm and other characteristics are concerned, amplify that region, and then amplify your ROI with the primers that you are trying to use now. more steps and more of a chance to introduce errors, but this approach may get you the product that you want.

I agree it looks like a primer problem and wonder if one primer has homology to a repeat sequence. To test this I would run a "pcr" with only one primer to see if that is where the multiple bands are coming from. Also try running a gradient of DMSO in your normal pcr from 1% to 6% in 1% increments whci often forces only the specific binding of the primers to work. Also if you can find some hot start taq in your lab give that a try.

• I agree with above commentators. There is also evidence of smearing
• Have you checked your DNA with other primers that have worked well with other DNA sample preps
• Conversely have you tried performing a PCR with these same primers on other DNA samples that have worked well with other primers in the past