Wanted: T7 Polymerase plasmid

Justin Powell Genetics jacp1 at mole.bio.cam.ac.uk
Thu Apr 20 06:09:04 EST 1995


I am trying to screen a cosmid expression library with inserts cloned
adjacent to a T7 promoter and I find I get much higher transfection
efficiencies with strains such as Stratagene Xl1-Blue MRA rather than the
strains BL21(DE3) or JM109(DE3) which have a chromosomally encoded T7
polymerase (even with the expression of this polymerase repressed).  I
believe this may be due to the Stratagene strains being deficient in the
MCR and MRR restriction systems as my insert is methylated - if so I could
get round the problem by amplifying the library in the Stratagene strain
and then transfecting the BL21(DE3).  I would however prefer to be able to 
express the T7 polymerase in the Stratagene strain.  

Does anybody have a plasmid encoding the T7 polymerase under control of an
inducible promoter?  Ideally this would be a plasmid which is not based on
the ColE1 origin or else contains the gene on an insert which would be
easy to subclone into such a plasmid.

Justin Powell
Genetics Dept.
Downing Street
Cambridge
CB2 3EH
England
 




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