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the DNA is not symmetrical

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#1 hslab



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Posted 06 November 2004 - 08:49 AM

hi, pcrman, I hapen to meet a strange thing, I am studying gene methylation with MSP and BSP with tumor tissues. In one of these samples, I got the MSP product successfully, and when I do sequencing with BSP, I found some CpG is methylated and some is not, to my surprised , the location with unmethylation is where the MSP primer is selected for. But the primer of MSP and the primer of BSP is not selected for the one strand of DNA modification, they aimed at different DNA stand with modification. So only one possibility can explain it, that is the DNA is not SYMMETRIC , in one stand , CG is methylated ; in the other stand, CG was unmethylated,

Do you agree with me?

#2 pcrman



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Posted 06 November 2004 - 09:13 AM

Are you sure your BSP and MSP primers were designed using different strand of DNA and you are not talking about upstrand and downstream primers?

DNA methylation is supposed to be symmetric on both strands. One possiblility that counts for the discrepancy is that your MSP primers are not specific enough to anneal only to methylated CpG sites, which is not uncommon especially if your PCR conditions are not optimized.

Edited by pcrman, 06 November 2004 - 09:15 AM.

#3 many



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Posted 08 December 2004 - 02:57 PM

I found the same thing. I input the complement sequence for picking up the primer for BSP. I check the sequences and found that all the modification happend in the G of the CpG, but not the C of the CpG.

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