PCR primers were used for sequencing, why didn’t they work? Could the problem be a degraded primer?
While degraded primer could be the culprit, another possibility is that your PCR primers are good for PCR reactions but not suitable for sequencing reactions. For sequencing primers, we recommend that they are about 18-24 bases in length with a Tm of 56-60 degrees. Longer PCR primers usually do not generate good results for sequencing reactions. Also, not all PCR primers work well as sequencing primers. If your PCR primer is not generating good sequencing results, we recommend that you design a nested primer to use as your sequencing primer.
