Specific-purpose plasmid cloning vectors II. Broad host range, high copy number, RSF 1010-derived vectors, and a host-vector system for gene cloning in Pseudomonas

• Published in: Gene Vol. 16; no. 1; pp. 237 - 247
• Main Authors: Bagdasarian, M., Lurz, R., Rückert, B., Franklin, F.C.H., Bagdasarian, M.M., Frey, J., Timmis, K.N.
• Format: Journal Article
• Language: English
• Published: Elsevier B.V 1981
• Subjects: bacteriophage λ; cosmids; Genetic engineering; mobilization-defective vectors; restriction endonucleases; soil bacteria; Km; cosmids; Cm; Hv; Tc; bacteriophage λ; restriction endonucleases; mobilization-defective vectors; kb; Genetic engineering; Sm; soil bacteria; Cb
• ISSN: 0378-1119; 1879-0038
• DOI: 10.1016/0378-1119(81)90080-9

Host-vector systems have been developed for gene cloning in the metabolically versatile bacterial genus Pseudomonas. They comprise restriction-negative host strains of Pseudomonas aeruginosa and P. putida and new cloning vectors derived from the high-copy-number, broad-host-range plasmid RSF1010, which are stably maintained in a wide range of Gram-negative bacteria. These plasmids contain EcoRI, SstI, HindIII, XmaI, XhoI, SalI, BamHI and ClaI insertion sites. All cloning sites, except for BamHI and ClaI, are located within antibiotic-resistance genes; insertional inactivation of these genes during hybrid plasmid formation provides a readily scored phenotypic change for the rapid identification of bacterial clones carrying such hybrids. One of the new vector plasmids is a cosmid that may be used for the selective cloning of large DNA fragments by in vitro λ packaging. An analogous series of vectors that are defective in their plasmid-mobilization function, and that exhibit a degree of biological containment comparable to that of current Escherichia coli vector plasmids, are also described.