This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Hosted, T. J. Articles by Baltz, R. H. Search for Related Content PubMed PubMed Citation Articles by Hosted, T. J. Articles by Baltz, R. H.

 Previous Article  |  Next Article 

J. Bacteriol., Jan 1997, 180-186, Vol 179, No. 1
Copyright © 1997, American Society for Microbiology

Use of rpsL for dominance selection and gene replacement in Streptomyces roseosporus

TJ Hosted and RH Baltz
Lilly Research Laboratories, A Division of Eli Lilly and Company, Indianapolis, Indiana 46285, USA.

We developed a gene replacement system using the rpsL gene of Streptomyces roseosporus and demonstrated its utility by constructing a deletion in the S. roseosporus glnA gene. A 1.3-kb BamHI fragment that hybridized to the Mycobacterium smegmatis rpsL gene was subcloned from an S. roseosporus cosmid library and sequenced. Plasmid pRHB514 containing the rpsL gene conferred streptomycin sensitivity (Sm(S)) to the Sm(r) S. roseosporus TH149. The temperature-sensitive plasmid pRHB543 containing rpsL and the S. roseosporus glnA gene disrupted with a hygromycin resistance (Hm(r)) gene was introduced into S. roseosporus TH149, and recombinants containing single and double crossovers were obtained after a temperature increase. Southern hybridization analysis revealed that single crossovers occurred in the glnA or rpsL genes and that double crossovers resulted in replacement of the chromosomal glnA gene with the disrupted glnA. Glutamine synthetase activity was undetectable in the recombinant containing the disrupted glnA gene.

This article has been cited by other articles:

• Dubeau, M.-P., Ghinet, M. G., Jacques, P.-E., Clermont, N., Beaulieu, C., Brzezinski, R. (2009). Cytosine Deaminase as a Negative Selection Marker for Gene Disruption and Replacement in the Genus Streptomyces and Other Actinobacteria. Appl. Environ. Microbiol. 75: 1211-1214 [Abstract] [Full Text]  
• van der Geize, R., de Jong, W., Hessels, G. I., Grommen, A. W. F., Jacobs, A. A. C., Dijkhuizen, L. (2008). A novel method to generate unmarked gene deletions in the intracellular pathogen Rhodococcus equi using 5-fluorocytosine conditional lethality. Nucleic Acids Res 36: e151-e151 [Abstract] [Full Text]  
• Doekel, S., Coeffet-Le Gal, M.-F., Gu, J.-Q., Chu, M., Baltz, R. H., Brian, P. (2008). Non-ribosomal peptide synthetase module fusions to produce derivatives of daptomycin in Streptomyces roseosporus. Microbiology 154: 2872-2880 [Abstract] [Full Text]  
• Gal, M.-F. C.-L., Thurston, L., Rich, P., Miao, V., Baltz, R. H. (2006). Complementation of daptomycin dptA and dptD deletion mutations in trans and production of hybrid lipopeptide antibiotics.. Microbiology 152: 2993-3001 [Abstract] [Full Text]  
• Miao, V., Coeffet-LeGal, M.-F., Brian, P., Brost, R., Penn, J., Whiting, A., Martin, S., Ford, R., Parr, I., Bouchard, M., Silva, C. J., Wrigley, S. K., Baltz, R. H. (2005). Daptomycin biosynthesis in Streptomyces roseosporus: cloning and analysis of the gene cluster and revision of peptide stereochemistry. Microbiology 151: 1507-1523 [Abstract] [Full Text]  
• Martinez, A., Kolvek, S. J., Yip, C. L. T., Hopke, J., Brown, K. A., MacNeil, I. A., Osburne, M. S. (2004). Genetically Modified Bacterial Strains and Novel Bacterial Artificial Chromosome Shuttle Vectors for Constructing Environmental Libraries and Detecting Heterologous Natural Products in Multiple Expression Hosts. Appl. Environ. Microbiol. 70: 2452-2463 [Abstract] [Full Text]  
• Sung, C. K., Li, H., Claverys, J. P., Morrison, D. A. (2001). An rpsL Cassette, Janus, for Gene Replacement through Negative Selection in Streptococcus pneumoniae. Appl. Environ. Microbiol. 67: 5190-5196 [Abstract] [Full Text]  
• Mchenney, M. A., Hosted, T. J., Dehoff, B. S., Rosteck, P. R. Jr., Baltz, R. H. (1998). Molecular Cloning and Physical Mapping of the Daptomycin Gene Cluster from Streptomyces roseosporus. J. Bacteriol. 180: 143-151 [Abstract] [Full Text]