Why probe hybridizes to vector?

faerber at ubaclu.unibas.ch faerber at ubaclu.unibas.ch
Tue Jul 11 09:54:30 EST 1995


In article <mic-1007952151470001 at 165.124.225.226>, mic at nwu.edu (Mic Chaudoir) writes:
> In article <biolc4-020795165223 at mac-918.sr2-building.uh.edu>,
> biolc4 at jetson.uh.edu (Wenfu) wrote:
> 
>> When I was using isolated DNA fragment( I am sure no any vector sequence in
>> it)
>> to probe digested plasmids, I found my probe hybridizes to the vector.
>> This kind of thing happen to me several times when I was doing the similar
>> thing. Although it did not matter for my results, I am really curious 
>> about the reasons for.
>> Any idea would be appreciated
> 
> Whenever you gel purify something, a residual amount of other DNA in the
> mixture comes along.  For example, if one cuts out a single band from PKS,
> and gel purifies it, some PKS comes along too.  Usually the amount is so
> small that it makes no difference.  However, the only way to completely
> eliminate it is multiple rounds of gel purification (actually, the
> contamination will still be there, just too small to be detectable by any
> means)........ 
> -- 
> Name:Mic Chaudoir    
> E-mail:mic at nwu.edu
> WWW:http://pubweb.acns.nwu.edu/~chaudoir/micpage.html
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Further comment:
for this reason we prefer to do a PCR with "insert-primers" and the insert as
the matrix. The product is really only insert without any traces of vector DNA.
This PCR product is then used as a template for i.e. Random Priming Labelling.

Good luck

Pit Faerber
Biozentrum Basel
faerber at ubaclu.unibas.ch

"That you're paranoid does not mean they are not out to get you."



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