Charged vs. uncharged nylon membranes?

Stephen Perry bafq5 at central.susx.ac.uk
Fri Nov 10 07:06:17 EST 1995


ssilverstone at csubak.edu wrote:
: A question:
: I'm dot-blotting total bacterial nucleic acids then probing with DNA
: probes complementary to the 16S rRNA. The protocol I'm looking at
: specifies uncharged nylon membranes, but the Amersham literature says that
: their Hybond N+ is the best membrane for RNA blotting (positively charged
: nylon). Does anybody know why charged or uncharged nylon might be
: preferred for probing RNA dot-blots with DNA probes?  I am planning to
: label the probes using the Amersham ECL kit.-Sara


Hi,

I have used N+ in the past, but always found there was a backgound
problem. For dot blots I would be particularly sceptical as you have
no separation of RNA species so you cannot judge whether the probe is
hybridizing specifically. I only used it for a couple of Northerns and
gave up when just about everything + the markers lit up!! I did not take
the time to alter the hybridization conditions or probe conc or washing,
so you may have more luck.

good luck  Steve Perry





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