methylation vary with passage number? - (Aug/01/2005 )

Hi,

I performed MSP on a cell line at passage 34 which showed both bands for M and U primers, so I assumed the gene is partially methylated. However, when I repeated the MSP with the same cell line but at passage 19, the M band disappeared (cell pellets were collected at different time). The DNA was not over-treated because another gene showed methylated using the same modified DNA. I wonder does methylation possibly vary with passage number? Thanks for any suggestion or comment.

It could be. But before you jump to that conclusion, verify it with bisulfite sequencing, which means you have to amplify the DNA using BSP primers.

There is a paper in the literature that describes this (Antequera et al 1995). In it, it describes how CpG islands on non-essential genes lose their ability to maintain their hypomethylated state and gain methylation over time. Thus from it, the authors state that results from methylation studies performed with derived cell-lines could have arisen from a culture artefact.

But do verify this by BSP as MSP only tests 1-3 CpG sites.

Good luck!!

Nick

pcrman and Nick,

Thanks for your suggestion.
Nick, could you please give me the details of that paper? I seem couldn't find it. Thanks again.

Whoops my bad, quoted the wrong year.

its :

Antequera F, Boyes J, Bird A. Cell. 1990 Aug 10;62(3):503-14.

entitled:

High levels of de novo methylation and altered chromatin structure at CpG islands in cell lines.


Nick