Biotin in hybridisation.

Dr. A. Rosenthal arosenth at hgmp.mrc.ac.uk
Tue Aug 30 10:01:31 EST 1994


Hallo Netters,
I am hybridising short hexamer labelled probes to a 2.5kb PCR biotinylated
target sequence at 60 celsius, but only about 1/3 of the probe binds to the 
target and only when the target is in 100 fold excess.
I was wondering if anyone had any experience with biotinylated probes in
hybridisation. What are the effects of randomly incorporated biotin nucleotides
like 16-dUTP  or 11-dUTP on the hybdridisation kinetics particulary when
hybridising in solution. Does the biotin lower the Tm?


Any comments are grately appreciated,

thanks.

Oliver Coutelle
Genome Analysis Group,
IMB Jena, German



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