Site-specific integration in Saccharopolyspora erythraea and multisite integration in Streptomyces lividans of actinomycete plasmid pSE101
Open Access
- 1 May 1988
- journal article
- research article
- Published by American Society for Microbiology in Journal of Bacteriology
- Vol. 170 (5) , 2287-2295
- https://doi.org/10.1128/jb.170.5.2287-2295.1988
Abstract
An 11.3-kilobase-pair plasmid, designated pSE101, exists in Saccharopolyspora erythraea NRRL 2338 as an integrated sequence (pSE101int) at a unique chromosomal location and in the free form in less than an average of 1 copy per 10 chromosomes. The plasmid sequence is missing from S. erythraea NRRL 2359. Restriction maps of the free and integrated forms of pSE101 showed point-to-point correspondence. Plasmid pECT2 was constructed by ligation of pSE101, pBR322, and the gene for thiostrepton resistance (tsr). When introduced by polyethylene glycol-mediated transformation into protoplasts of S. erythraea NRRL 2359, all thiostrepton-resistant regenerants examined were found to carry a single copy of pECT2 in the integrated state at a single chromosomal site. The chromosomal site of pECT2 integration in strain NRRL 2359 (attB) corresponded to the chromosomal location of pSE101int in strain NRRL 2338. The plasmid crossover site (attP) was mapped to the plasmid site that corresponded to the site of interruption of the plasmid sequence in the host carrying pSE101int, indicating that site-specific integrative recombination had occurred. An additional 2.8-kilobase-pair chromosomal sequence homologous to a segment of pSE101 was also observed in strains NRRL 2338 and NRRL 2359. After introduction of pECT2 into Streptomyces lividans, approximately half of the transformants examined were found to carry the plasmid as a stable, autonomously replicating element. The other half carried a single copy of pECT2 as an integrated sequence, but the location of pECT2int in Streptomyces lividans varied from one transformant to another. In each case, integrative crossover used the attP site. A model is proposed to account for the determination of the particular state of pSE101 in Streptomyces lividans.This publication has 27 references indexed in Scilit:
- Detection of specific sequences among DNA fragments separated by gel electrophoresisPublished by Elsevier ,2006
- Isolation and characterization of an extrachromosomal element from Nocardia mediterraneiPublished by Elsevier ,2004
- Excision and integration of a self-transmissible replicon of Streptomyces ambofaciensGene, 1987
- Transfer of the Type Strain of Streptomyces erythraeus (Waksman 1923) Waksman and Henrici 1948 to the Genus Saccharopolyspora Lacey and Goodfellow 1975 as Saccharopolyspora erythraea sp. nov., and Designation of a Neotype Strain for Streptomyces erythraeusInternational Journal of Systematic and Evolutionary Microbiology, 1987
- Cloning and Expression of the Tyrosinase Gene from Streptomyces antibioticus in Streptomyces lividansMicrobiology, 1983
- Plasmids, Recombination and Chromosome Mapping in Streptomyces lividans 66Microbiology, 1983
- Construction and characterization of new cloning vehicles III. Derivatives of plasmid pBR322 carrying unique Eco RI sites for selection of Eco RI generated recombinant DNA moleculesGene, 1978
- Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase IJournal of Molecular Biology, 1977
- A complementation analysis of the restriction and modification of DNA in Escherichia coliJournal of Molecular Biology, 1969
- A membrane-filter technique for the detection of complementary DNABiochemical and Biophysical Research Communications, 1966