I run PCR to checked presence of specific gene in shigella but I received 2 bands, one band showed product size expected and another one showed higher product size. I would like to check the my primer pair may have multiple binding sites or not. I did BLAST primer and got the result with 1 primer pair. How do I know that my primer pair has multiple binding sites through this result? Any other websites can check whether the exact position that my primers bind to the genome.
Your answers are really appreciated.
Can you share a picture of your gel? How bright is the unexpectedly large band? Does it show up in your negative control?
One easy first thing to try is use a gradient of annealing temperatures. That might be enough to get your PCR condition specific to your desired product.
Good luck!
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