SS Nytran-- Please Help!

Daniel Meier dmeier at post.its.mcw.edu
Mon Nov 21 11:39:59 EST 1994


Dear All,

  Over the past years I have successfully used Schleicher & Schuell Nytran.
About a month ago, our supplies ran out and we purchased additional SS
Nytran.  Since our previous purchase, Scleicher & Schuell changed the
formulation of their Nytran (apparently due to patent reasons).  When we use
this new Nytran with our old hybridization conditions we have significantly
high backgrounds (very dark at 3 days exposure).
  We hybridize at 42 C with 50% formamide, 25 mM KOP4 (pH 7.4), 5x
Denhardt's, 50 um/ml herring sperm DNA, and 5x SSC.  We wash blots 2x at RT
for 15 min each using 2X SSC,0.1% SDS and 2x 15 min each using 0.25X SSC,
0.1% SDS at 57C.  Our probe was gel-purified from a 1.5% Boehringer-Mannheim
agarose gel.  Any suggestions how we might lower our background using these
membranes?  Can you suggest a membrane that works well for you and allows
multiple stripping and reprobing?

Thanks much,

Dan Meier
dmeier at post.its.mcw.edu




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