Panomics Promoter Methylation PCR Kit and Methylation specific Digests - (Aug/05/2008 )
I'm trying to determine the methylation status of my promoter of interest and would love some advice. I've tried doing bisulfite sequencing which was inconclusive, so am taking a step back to first determine if in fact my promoter is methylated. I wanted to ask if anyone has used the promoter methylation PCR kit from panomics and if they have had success.
I also want to simply run a digest (to determine if there is a genomic methylation difference between cell lines) with methylation specific or sensitive res. enzymes (which would be better?) on genomic DNA and was hoping someone could give advice on which enzymes to use and the order in which to use them (ie should I cut the genomic DNA isolated from the cell line (I have a cancer cell line and a wild type I will be using as a control) with an ezyme to make smaller fragments followed by methylation specific or sensitive enzymes?).
Any help would be appreciated,
why not try qMSP method for interested targets? This is a recent reference at http://cancerres.aacrjournals.org/cgi/cont...ull/67/21/10117.
what do you mean with BSP was inconclusive - in my opinion this is the best way to determine (a) the amount of methylation if ( methylation is present in your promoter of interest.
Restriction enzymes can be used, but I don't trust them too much. We used HpaII and MSPI.