pGEX fusion vector expression

GIETZ at bldghsc.lan1.umanitoba.ca GIETZ at bldghsc.lan1.umanitoba.ca
Mon Jan 25 13:49:00 EST 1993


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I have just cloned my gene into the pGEX vector using XL1-blue
cells.  Now the question is this:  is XL1-blue a suitable cell
line for expressing my fusion protein?  If not, what is a good
cell line to use?  Anyone who have had experience with the pGEX
vector please respond.

-fm

ps.  pGEX is a glutathione S-transferase fusion vector.
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-fm
    We have used pGEX successfully for a number of genes
and have found that JM109 is the best strain for the production
of soluable fusions when compared to DH5alpha.  There was about
a 5-10 fold better production of full length fusion in JM109.  We have
also found that specific growth media also helps increase
the yield of active (properly folded?) protein.  
Blackwell and Horgan (1991) Febs letts 295;p10-12(Thanks to Kiran Madura)
describes a method using betaine in LB media with sorbitol to increase 
the yield of active( properly folded) fusion proteins.  We have tried it and 
it works very well for both our fusion proteins.

________________________________________
Dan Gietz
Department of Human Genetics
University of Manitoba
770 Bannatyne Ave, Rm 250
Winnipeg, Man, Canada
R3E 0W3
GIETZ at BLDGHSC.LAN1.UMANITOBA.CA
_________________________________________




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