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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