Mike Chao mikechao at hamon.swmed.edu
Thu Jan 16 15:40:01 EST 1997

In article <33EC032025 at puskin.sote.hu>, SZABADKAI at PUSKIN.SOTE.HU
("Szabadkai Gyorgy") wrote:

>I tried to use Hybond N membranes for dot blots of RNA and DNA, but 
>the DNA signal has lost somehow. I denaturated the DNA by boiling for 
>5 minutes and then fixed on the membrane by UV crosslinking (1200 
>mJ/cm2). The probe was ok, I checked it.
>What kind of denaturation and fixation do you use for this memebrane?
>Szabadkai Gyorgy

for denaturing we use akaline (i.e., the standard NaOH/NaCl solution for
most Southerns), UV X-linking for fixation. For Hybond we found that if you
wash the membrane with clean H2O after X-linking and just before adding hyb
buffer, this reduces background.


Mike Chao                                       
UTSW Medical Center @ Dallas
Dept. Mol. Biol. & Oncology
Phone: (214) 648-1489
Fax:   (214) 648-1488

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