Hybridization oven question

LAZO,GERARD glazo at aardvark.ucs.uoknor.edu
Mon Jun 28 08:54:00 EST 1993


>We have just bought a hybridisation oven and i was wondering if you all out
>there have any tips on making the most effective use out of it..
>
>What has been the maximum number of filters used per bottle?
>> I have used up to 5

I often probe microtiter dish formatted filters and have used 
up to 15 filters per tube for a single hybridization.

>Do you really need to use the mesh provided for placing between filters?
>> Dunno but always have.

I've never used this mesh before.  Since I have been using so many 
filters per tube I might consider this.  What kind of mesh are we
talking about here?  Is it a special product?

>What kind of hyb solution volume to surface area of filter do people use?
>> Varies for target and probe i.e DNA-DNA, DNA-RNA etc

I use 20 ml in a tube with 15 filters.

>Has anyone altered the hyb soln composition as a result of using these ovens?
>> No

No, but I use the Church procedure.  However, I have had some
problems with stripped and re-probed filters.  For some reason
the newly-labelled probe has an affinity for some of the microtiter
dish wells which gave me a positive signal on a previous hybridization.

>Are any brands of filter/membrane better than others in these ovens.?
>> I use Hybond N+ and alkaline transfer.

I use ZetaProbe filters.

Gerard R. Lazo               /\     //====== glazo at aardvark.ucs.uoknor.edu
Plant Biology Division      // \   //            LAB: 405-221-7305
The S.R. Noble Foundation, Inc. \ //======           FAX: 405-221-7380
Ardmore, Oklahoma 73402   //     \/                      MSG: 405-223-5810



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