Electropor./conjug. in Pediococcus

Hank Seifert h-seifert at nwu.edu
Fri Apr 29 09:48:54 EST 1994


In article <2pqimb$71n at maya.dei.unipd.it> alessio at civ.bio.unipd.it (Giacomini Alessio) writes:
>In our lab we are working on a strain of Pediococcus pentosaceus, a
>Gram + lactic acid bacterium. We need to transfer a broad host range
>vector into it. So far we have tried both electroporation and conjugation
>(from E.coli S17-1) but without result. I know from published work that
>that these bacteria seem to be somewhat recalcitrant to these procedures,
>and transfer frequencies are always very low (in not equal to zero, as
>in my case!). Does anyone know a good protocol or can give an advice to
>overcome this problem?
>Thank you VERY much
>
>Alessio Giacomini
>CRIBI Biotechnology Centre
>Univ. Padova - ITALY
>E-mail: alessio at cribi1.bio.unipd.it
Have you considered that your problem is restriction?  If it 
is, do a conjugation or electroporation, isolate the 
transconjugants/transformants, cure the plasmid, and retest to 
see if you get higher frequencies.  Often the few isolated in 
the first round have inactivated the restriction activity.
Just a thought!
Hank Seifert
Northwestern University



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