Oligo hybridization.

Harold J. Drabkin hdrabkin at user1.channel1.com
Tue Jan 27 11:53:14 EST 1998


On Fri, 23 Jan 1998 12:18:59 +0100, Peter Nilsson
<Peter.Nilsson at klinkemi.umu.se> wrote:

>Do anyone have a fast and easy protocol for blotting and hybridization of PCR products (100-400 bp) using end-labelled oligonucleotides?
>

I might be tempted to use a 3-5 % native acrylamide gel (1:29 x-link,
TBE buffer), and transfer to Nytran +.  Hybridization to probles of
18-30 is as follows:
Prehyb: 42C in 10X Denh., 6XSSC, 1% SDS (hybrid. oven recommended).
for 2 hrs.
Hyp: 6XSSC, 1% SDS at 42C overnight
Wash:
2x 10 min in 6xSSC
2X 10 min in 2X SSC
Background usually not a problem; you might be able to increase
stringency with 1x or 0.5x SSC.




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