Bacterial expression of reverse-frame plasmid genes

mbraden23 at mbraden23 at
Tue Oct 31 16:38:31 EST 2000


I am in the process of subcloning lacZ into a plasmid made for mammalian
expression.  With the restriction enzymes Ive used, the gene should be
inserted in frame with the mammalian promoter (CMV).  This is reverse of
the reading frame for the bacterial promoters/genes, such as the
ampicillin resistance.  I was wondering if the bacteria (JM109) would
still be able to express the lacZ gene, thus providing additional
selection for proper insertion/ligation (such as growth on X-Gal/IPTG
plates).  Would it just be easier to have the ligated plasmid sequenced
to verify the framing of the insert?

Mike Braden
Marker Gene Technologies

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