BRL RNA ladder hybridizes to my probe

Steven Poet poets at ccmail.orst.edu
Thu Aug 17 17:56:43 EST 1995


You've got vector sequence contamination.  Try generating your probe 
with internal PCR primers.  BRL's got a PCR labelling kit which works
well.  Use a very, very, very small amount of template in the reaction.


stallard at badlands.NoDak.edu (Lana C Stallard) wrote:
>
> I wonder if anyone has had this happen? I am using Gibco BRL RNA weight 
> standards (made from phage lambda DNA spliced to an RNA transcript). I get 
> significant hybridization with a mouse DNA probe (a cDNA probe of 
> angiotensin converting enzyme) to the ladder when i transfer the ladder 
> onto a Northern blot. Why would there be so much sequence similarity between lambda and a mouse gene? or could my probe still contain some sequences of the vector it was produces in (Bluescript 
> plasmid) which is similar to the phage DNA? My boss is convinced i have a 
> very non-specific probe but i think there must be a reason. 
> 	Thanks for any comments or ideas you may have. 
> 	Lana Stallard




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