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 (RSF 1010), (ii) IncQ-Iike pTF-FC2 and (iii) pSB 102 that belongs to a new class of broad-host-range plasmids. The established system based on the IncP(x 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 IncP alpha-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. (c) 2005 Elsevier B.V. All rights reserved.