Transfer and establishment of DNA in Streptomyces (a brief review).
Based on endogenous Streptomyces plasmids we have constructed various multiple purpose vectors for cloning in streptomycetes. Since replication of the S. ghanaensis plasmid pSG5 is inherently temperature-sensitive, pGM-vectors derived from pSG5 can be used for gene disruption/replacement and mutational cloning. The 1.6-kb minimal replication region of pSG5 encodes only one polypeptide, an initiation protein (Rep) for single stranded DNA-replication. Different internal fragments of sequenced genes of the Ptt-biosynthetic pathway were cloned into pGM-vectors to study both, the function of the biosynthetic genes and recombination in Streptomyces. The observed recombination frequencies were very high, up to 80% of the cells carried single or multiple copies of the plasmid integrated into the chromosome. Replacement experiments revealed that the frequency of marker exchange and plasmid integration, respectively, lies in the same order of magnitude. The region of homology which is required for homologous recombination could at least be reduced to 200 bp.
Wohlleben
W
W
Institute of Genetics, Biological Faculty, Bielefeld University, Germany.
Hartmann
V
V
Hillemann
D
D
Krey
K
K
Muth
G
G
Nussbaumer
B
B
Pelzer
S
S
D003001:Cloning, Molecular / Q000379:methods; D004269:DNA, Bacterial / Q000235:genetics*; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic; D013302:Streptomyces / Q000235:genetics*
D004269:DNA, Bacterial