omega fragment of Beta-galactosidase

Michael Benedik bchs1b at Elroy.UH.EDU
Thu Mar 10 16:36:56 EST 1994


In article <Jonathan_Kurtis-090394180651 at tonto-slip6.cis.brown.edu>, Jonathan_Kurtis at Brown.edu (Jonathan Kurtis) writes:
>I tried mailing this question to Stratagene's new Tech Service email
>account, but have not gotten a response. Could anyone out there please
>answer the following questions:
>
>1) does the lacZÆM15 gene on the F' episome in SOLR and XL1-Blue MRF' cells
>code for the omega fragment of Beta-galactosidase ??
>
>2) if it does, how big is this protein (in Kda) ?
>
>3) is it constituativly expressed or is it induced by IPTG?
>
>The reason for these questions is as follows: I immunoscreened (with a Mab)
>a Lambda ZAP II library and plaque purified a positive clone. I have
>excised the pBluescript SK plasmid into SOLR. When I induce a culture of
>these cells, make an E. coli lysate, and run it out on a gel, I get an
>induced band at 30 Kda that is recognized by my Mab (this is consistent
>with the size of my cDNA insert =1.7 kilobases with some 3' untranslated
>region). I also get an induced band at around 100 kDa (too big to be coded
>for by my insert) that is not recognized by my Mab. I thought that this
>band might be due to the induction of the omega fragment of B-gal.
>
>Thanks for your help, 
>
>
>-- 
>Jonathan Kurtis
>
>International Health Institute
>Brown University Box G-8033
>Providence, RI  02912



The answers are Yes, Yes and Yes.
The lacZdeltaM15 allele encodes the omega fragment of lacZ, which is
essentially is the entire lacZ except for a small in-frame deletion
near the beginning. It should  be about 100-110 kDA. It should also be
inducible, but if there is a high copy plasmid around it will probably be
expressed without IPTG, just maybe not maximally.


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 Michael Benedik				INTERNET: Benedik at uh.edu
 Dept. of Biochemical & Biophysical Sciences	
 University of Houston				BITNET: Benedik at uhou
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