checking for DNA on nylon membranes?

Casey M. Finnerty cmf5 at cornell.edu
Mon Apr 18 15:24:34 EST 1994


In article <rcaspi.14.2DADD381 at ucsd.edu>, rcaspi at ucsd.edu (Ron Caspi)
wrote:

> I have some nylon membranes (HiBond N) that once had DNA on them (transferred 
> by southern blotting). These membranes have been hybridized and stripped, so 
> the DNA is now single stranded. However, my last probing yielded nothing - no 
> signal whatsoever. I suspect that I might have lost the DNA.
> Does anybody know how I could check if the DNA is still there?

You can try staining with 0.02% methylene blue in 0.3M NaOAc, pH 5.5 at
room temp. for a few minutes. This stain is reversible and does not
interfere with hybridization. If you wish to destain, shake the blot in
0.2X SSC, 1% SDS at room temp. The technique works better for RNA, but it
will also detect DNA. It is described in Maniatis for RNA (at least the old
manual) and I'm told it's also in Biotechniques (1988) Vol 6 No 3, pp
196-8.
-- 
Casey M. Finnerty
Boyce Thompson Institute at Cornell University
cmf5 at cornell.edu



More information about the Methods mailing list