Riboprobes, Northerns, and high background

slavemaster z un691cs at genius.embnet.dkfz-heidelberg.de
Mon Nov 25 11:16:04 EST 1996


In article <tyr-2-2011961020250001 at news.srv.ualberta.ca>, tyr-2 at bones.biochem.ualberta.ca 
says...
>
>In article <56v7mj$23ic at r02n01.cac.psu.edu>, kgilbert at cmp.hmc.psu.edu
>(Gil) wrote:
>
><snip> 
>> I detect single bands on Northerns, but get high lane background.  Any
>> suggestions on ways to eliminate this high backgorund would be greatly
>> appreciated.  Thanks in advance.
>
>One thing we've found (and other have posted similarly) that the blocking
>components of your hyb solution can change depending on the type/brand of
>membrane you use. One such membrane distributer suggests increasing SDS to
>1 % final while another retains SDS at 0.1% (standard for our Southerns)
>yet increases Denhardt's solution to 10X final concentration rather than
>5X.
>
>Bottom line - take a look at the distributer's recommendations for the
>membrane (a la Schleicher and Schuell's Product Guide and Methods - no
>affiliation). If there's nothing in the catalogue or accompanying the
>product call their tech rep and they will usually fax you the info on the
>same day.

Yep, but strangely enough a hyb buffer with 7 % (yes: seven percent !) SDS
and NO denhardt's results in very low background with practically all filters
(Nitrocellulose, genescreen, Hybond N and N+, Zetaprobe).

Try it ! It works wonders. For exact buffer conditions, send an e-mail to:

slavemaster z

at the address indicated above.




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