Hi Background Plague Lifts

Ken Soderstrom soderstk at ucs.orst.edu
Thu Oct 9 13:17:13 EST 1997


Are you making your probe by random priming?  What is the source of the

The one case of all-plaque-labeling that I'm aware of was due to the
template used for random primer probe synthesis.  This template was a PCR
fragment generated with vector primers that contained about 50 bp of vector
sequence.  The entire fragment was ~800 bp but those 50 vector bases, under
probably not too stringent conditions, was enough to label every plaque.

Hope you solve your problem,

Ken Soderstrom
soderstk at ucs.orst.edu

Michael Salvucci <mesalvu at ix.netcom.com> wrote in article
<61j49b$rho at dfw-ixnews8.ix.netcom.com>...
> Dear Netters,
> We are having a problem with very high backgrounds in our plaque lifts.  
> problem is that all of the plaques are hybridizing with our probe.  The
> problem occurs with both DNA and riboprobes for different genes, even
> we are using standard and even strigent conditions for prehybridization,
> hybridization and washing.  We are using a library prepared in the
> Express phagemid and infecting a bacterial lawn of XL-1 Blue cells.  Any
> suggestions would be greatly appreciated.  Please email me directly. 
> Thanks........MIKE

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