Retrieving DNA from lysed cell pellet
Rob Jordan
rjordan at u.washington.edu
Tue Sep 24 13:55:42 EST 1996
I'm working on a project where I am irradiating cells and isolating
low molecular weight DNA at various times after irradiation. To do
that I'm using a method similar to one I use to isolate DNA from cells
undergoing apoptosis: lyse cells in 0.6% SDS, 10 mM EDTA then add
NaCl to 1 M. Incubate overnight at 4C then spin down the goo.
Presumably the supernatent contains low molecular weight DNA. It's
basically the Hirt method.
Anyway as a sort of control I want to get at the DNA that's in pellet.
What's the best way to do that? I was thinking of suspending the
pellet as best I could in something, treating with proteinase, then
phenol-chloroform extracting. I don't need a huge amount of DNA and I
don't need it to be a particular size. Is this a reasonable way to do
it? If so what should I resuspend the pellet in? I was thinking
maybe 0.2X SSC. But I really have no idea what would be best.
Thanks-
Rob
rjordan at u.washington.edu
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