bisulfite modification after pcr - (Aug/16/2005 )
has anyone ever tried doing a bisulfite modification on a pcr product? i am wondering if this will increase the specificity of my final pcr product when i do a msp after the modification. if this makes sense...
1. pcr of promoter region that encompasses my msp primers
2. bisulfite modification
3. mspcr (like a nested pcr reaction)
is this doable?
Short answer, no, it is not possible. The PCR reaction incorporates unmethylated dNTPs into the product strands. Any information about the methylation state of the original template strand is destroyed in that process. The bisulfite reaction following the PCR will convert all of the C's to T's, regardless of the methylation status of the original genomic DNA.
Regular PCR cannot tell the difference between C and mC, so all of the PCR products will be same, this means you can not get methylated products....