expression vectors for proteins

Andrea Skantar seh at bchm.biochem.duke.edu
Wed Sep 16 15:00:56 EST 1992


In article <9209130602.AA26308 at schooner.waite.adelaide.edu.au>, rbonfigl at waite.adelaide.edu.au (Rod Bonfiglioli) writes:
> 
> does anyone have any experience of using expression vectors for largish
> proteins, i am trying tofind one that will take inserts of up to 3 Kb to
> express proteins of the 100 Kd range.  i have tried pGEX vectors, and they


> 
Try the beta-galactosidase vector series pUR 290, 291, 292. They worked pretty
well for my subcloning of a 2kb fragment of the second-largest subunit
of Drosophila RNA polymerase II.  This construct was not very soluble, but I
was able to solublize the inclusion bodies in 8M urea + DTT followed by dialysis
into buffer with no salt in  it.  Since the pol II subunit is "used to" being
assembled next to a 215 kd protein and several other small subunits, the 
domain seemed to be "happier" next to something large, ie. beta-gal.  It's
a tetramer, but it might work for some applications.  Hope this helps.

Andrea



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