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mC not in CpG? Primer selection

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#1 sabo



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Posted 13 January 2005 - 12:35 PM

we are looking for 2 primer-pairs for nested PCR and subsequent sequencing of bisulfit-treated DNA. In case of primers with internal CpG dinucleotides, we have to design a mix of primers with C or T at the corresponding position. But what about the other C-residues within the primer sequence? Petronis et al. (Schizophr Bull 29:169ff (2003)) found in their study some methylated C-residues, which are not in CpG or CpNpG. Isn't there the danger of loosing templates and shifting of results?
Is there anybody who has experience with this problem?

#2 pcrman



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Posted 13 January 2005 - 01:08 PM

For non-CpG 'T' in the primer, treat it as 'T'. Although there are studies reporting non-CpG methylation in mammlian systems, this is not well established and likely artfact due to incomplete conversion.

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