DNA from somatic tissues for bisulfite treatment - (Dec/06/2006 )
I am in a lab that does a fairly significant amount of methylation analysis. However, we usually look at sperm DNA and not DNA from somatic tissues. Purifying DNA from the sperm is quite easy, considering there is a smaller amount of histones associated with the chromatin. So, we are having a little bit of trouble getting good clean DNA from the somatic tissues for the bisulfite conversion. Does anybody have a good protocol or tips for us?
a standard phenol/chloroform extraction works fine for bisulfite studies.
it is important to get rid of all proteins that are associated with the DNA. The following protocol works fine: http://methdb.igh.cnrs.fr/cgrunau/methods/...c_DNA_prep.html
yes, it's also good to keep good histological data for somatic studies.
I've done tissue work from many sources. Some of the downstream research involves restriction enzymes, so we do not use phenol methods as they inhibit digests. I highly recommend the Epicentre kit. Very clean DNA for multiple downstream methods.