We realized we'd been using a restriction enzyme (to digest our PCR products for genotyping) which is sensitive to CpG methylation. We are now worried that this will affect our results, and we have to do everything all over again. In literature, that particular RE is used repeatedly, and none of the articles mentions about the possible effect of methylation.
Thank you for your help!
Even if your enzyme is sensitive to methylation, you don't have to worry. Your PCR product lacks methylation. If you check your genotype by southern blotting CG methylation is again a topic.
Uwe is right, regarding lack of methylation in the PCR amplicons. However, if you are focused at examining the methylation status at certain locus of DNA, you have to carefully select your REs. You have to re-design your detection method, by doing RE digestion of the native genomic DNA, followed by PCR amplification.
SD
Both answers are right. PCR products are not methylated. We also had some experiments where we analyzed CpG methylation patterns within PCR products. First, we had to enzymatically methylate theses DNA strands, since they were not methylated directly after the PCR.
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