Betalactamase assay or copy number estimate needed

Bernard P. Murray, PhD bpmurray*STUFFER* at socrates.ucsf.edu
Wed Sep 16 18:31:39 EST 1998


In article <6tp9tk$723$1 at newsflash.concordia.ca>, neweb at alcor.concordia.ca
( ELAINE B S NEWMAN ) wrote:

> Help Needed- how to assay beta-lactamase
> We want to be able to estimate copy number of E. coli plasmid
> pBR322 by looking at the amount of beta-lactamase-
> however we don't know how to do that- assays either are complex
> and require a radioactive substrate,
> or they required a substrate which is not easily  available and
> very expensive ($1000/gm) when it is. 
 
> Does anyone have a good way to estimate beta=lactamase, or
> a good way to estimate copy number?
 
> If answers are sent directly to me (Neweb@ Vax2.concordia.ca}
> I will post a summary of them to this group. 
> Elaine Newman

Is there any reason that you want to avoid a Southern blot?
Alternatively, just performing a plasmid preparation and
checking the yield would give you a crude estimate of the
copy number.
If you want a comparative estimate you could simply
determine the resistance (eg. by MIC) of the bacteria
to ampicillin and/or tetracycline.
     Bernard
-- 
Bernard P. Murray, PhD
Dept. Cell. Mol. Pharmacol., UCSF, San Francisco, USA



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