Quick/Express/Easy/Etc Hybridization buffers?

Jean Harris jean.harris at stonebow.otago.ac.nz
Mon Oct 11 18:42:45 EST 1999


This message is in reply to:

________________
From: John Dixon <jpcd100 at mole.bio.cam.ac.uk>
Subject: Quick/Express/Easy/Etc Hybridization buffers?
Date: Thu, 07 Oct 1999 12:12:09 +0100

Hi All, can anyone give me any tips as to the true efficacy of all
these fancy hybridization buffers being advertised recently. Ambion had
an advert in Nature last week showing a northern hybed in "standard"
buffer and the same blot using their buffer, showing a massive increase
in signal.

Are any really as good as they say they are? Thanks for any info or
recommendations.

Cheers

John

-- 
John Dixon                    Lab 44 (1223) 334131
Wellcome/CRC Institute        Fax 44 (1223) 334089
Cambridge University
United Kingdom CB2 1QR       e-m: jpcd100 at mole.bio.cam.ac.uk
_________________

Dear John,

I also wonder what the active ingredients are whenever biotech 
companies advertise some fancy new reagent.  The 
Quick/Express/Easy/Etc Hybridization buffers greatly intrigued me. 
The magic ingredient is probably CTAB .  While pursuing a hobby 
interest in CTAB I came across these papers:

1.  Kumar A, Wilson SW.  (1990)  Studies of the Strand-Annealing 
Activity of Mammalian hnRNP Complex Protein A1,  Biochemistry 29; 
10717-10722.

2.  Pontius BW, Berg P.  (1990)  Renaturation of complementary DNA 
strands mediated by purified mammalian heterogeneous nuclear 
ribonucleoprotein A1 protein: Implications for a mechanism for rapid 
molecular assembly.  PNAS 87; 8403-8407.

3.   Pontius BW, Berg P.  (1991)  Rapid renaturation of complementary 
DNA strands mediated by cationic detergents: A role for 
high-probability binding domains in enhancing the kinetics of 
molecular assembly processes.  PNAS 88; 8237-8241.

The implication of these papers is that CTAB or A1hnRNP is the key 
ingredient in Boehringher Mannheim's Quick Hyb and the rapid 
hybridization solutions sold by other biotech companies.  CTAB is 
more likely since it is probably a lot cheaper than A1hnRNP and gives 
the same result.

The authors of refs 2 and 3 didn't indulge in much discussion of the 
obvious practical applications of their results.  That led me to 
speculate that they may have obtained a commercial spin-off from 
their work and didn't want to be too specific about how to apply it. 
The question is how much CTAB to chuck into a hyb solution and would 
it be worth playing around a bit with CTAB in hyb solutions to get a 
good formulation and avoid the terrific expense of the pre-made hyb 
solutions sold by various biotech companies.

I haven't used Southerns  for quite awhile myself so I have no 
motivation to try this at present.  If any of you out there were to 
do it, it should make a nice little paper in BioTechniques and pull 
the rug out from under the biotech companies as far as quick hyb-type 
solutions are concerned:)

Cheers,
Jean

PS  If anybody does such a paper I would appreciate acknowledgement 
for kicking it off.

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Eugenie L Harris, PhD		jean.harris at stonebow.otago.ac.nz
Research Fellow
Vascular Biology Group
Surgery Department			Phone:  +64 3 474-0999  ext. 7474
University of Otago			Fax:  +64 3 474-7622
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