The streptomyces genome contains multiple pseudo-attB sites for the (phi)C31-encoded site-specific recombination system.
The integrase from the Streptomyces phage (phi)C31 is a member of the serine recombinase family of site-specific recombinases and is fundamentally different from that of lambda or its relatives. Moreover, (phi)C31 int/attP is used widely as an essential component of integration vectors (such as pSET152) employed in the genetic analysis of Streptomyces species. phiC31 or integrating plasmids containing int/attP have been shown previously to integrate at a locus, attB, in the chromosome. The DNA sequences of the attB sites of various Streptomyces species revealed nonconserved positions. In particular, the crossover site was narrowed to the sequence 5'TT present in both attP and attB. Strains of Streptomyces coelicolor and S. lividans were constructed with a deletion of the attB site ((Delta)attB), and pSET152 was introduced into these strains by conjugation. Thus, secondary or pseudo-attB sites were identified by Southern blotting and after rescue of plasmids containing DNA flanking the insertion sites from the chromosome. The sequences of the integration sites had similarity to those of attB. Analysis of the insertions of pSET152 into both attB(+) and (Delta)attB strains indicated that this plasmid can integrate at several loci via independent recombination events within a transconjugant.
Combes
Patricia
P
Institute of Genetics, University of Nottingham, Queens Medical Centre, Nottingham NG7 2UH, United Kingdom.
Till
Rob
R
Bee
Sally
S
Smith
Margaret C M
MC
D001287:Attachment Sites, Microbiological / Q000235:genetics*; D001435:Bacteriophages / Q000235:genetics*; D001483:Base Sequence; D003227:Conjugation, Genetic; D017353:Gene Deletion; D014644:Genetic Variation; D016680:Genome, Bacterial; D019426:Integrases / Q000235:genetics; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic; D017422:Sequence Analysis, DNA; D013302:Streptomyces / Q000145:classification / Q000235:genetics* / Q000821:virology
D019426:Integrases