Introduction of DNA into Actinomycetes by bacterial conjugation from E. coli--an evaluation of various transfer systems.
Gene transfer is a basic requirement for optimizing bioactive natural substances produced by an increasing number of industrially used microorganisms. We have analyzed quantitatively horizontal gene transfer from Escherichia coli to Actinomycetes. The efficiencies of DNA transfer of four different systems were compared that consist of conjugative and mobilizable plasmids with a broad-host range. Three novel binary vector set-ups were constructed based on: (i) the IncQ group of mobilizable plasmids (RSF1010), (ii) IncQ-like pTF-FC2 and (iii) pSB102 that belongs to a new class of broad-host-range plasmids. The established system based on the IncPalpha group of conjugative plasmids served as the reference. For all plasmids constructed, we confirmed the functional integrity of the selected transfer machineries by intrageneric matings between E. coli strains. We demonstrate that the transfer systems introduced in this study are efficient in mediating gene transfer from E. coli to Actinomycetes and are possible alternatives for gene transfer into Actinomycetes for which the IncPalpha-based transfer system is not applicable. The use of plasmids that integrate into the recipients' chromosomes compared to that of plasmids replicating autonomously is shown to allow the access to a wider range of hosts.
Blaesing
Franca
F
Max-Planck-Institut für Molekulare Genetik, Ihnestrasse 63-73, D-14195 Berlin, Germany.
Mühlenweg
Agnes
A
Vierling
Silke
S
Ziegelin
Günter
G
Pelzer
Stefan
S
Lanka
Erich
E
D039903:Actinobacteria / Q000235:genetics*; D001483:Base Sequence; D001709:Biotechnology; D003001:Cloning, Molecular; D003227:Conjugation, Genetic; D004274:DNA, Recombinant / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics*; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D010957:Plasmids / Q000235:genetics; D013302:Streptomyces / Q000235:genetics
D004274:DNA, Recombinant