RRC ID 30046
Author Hashimoto-Gotoh T, Franklin FC, Nordheim A, Timmis KN.
Title Specific-purpose plasmid cloning vectors. I. Low copy number, temperature-sensitive, mobilization-defective pSC101-derived containment vectors.
Journal Gene
Abstract Two cloning vector plasmids, pHSG415 (7100 bp) and a lambda phage cos site-containing derivative (cosmid) thereof, pHSG422 (8760 bp), were constructed from a low copy number plasmid (pSC101) replicon to permit the propagation of cloned DNA segments at low gene dosage levels. Two features of the vectors, namely temperature sensitivity of replication and inability to be mobilized by conjugative plasmids, cause them to exhibit a high level of "biological containment". The essential characteristics of pHSG415 and pHSG422 may be summarized as follows: (1) their genome copy number is low (4--6 copies/chromosome); (2) their replication ceases at high temperature and they are rapidly lost from host cells grown at temperatures of 37 degrees C and above; (3) the relaxation nick site of pSC101, which is thought to be synonymous with its origin of transfer replication, is absent from the vectors; as a consequence, they are not mobilized to a significant extent by co-existing conjugative plasmids that are able to mobilize wild-type pSC101; (4) they contain unique insertion sites for DNA fragments generated by the following restriction endonucleases: EcoRI, XhoI, XmaI, HindIII and PstI; pHSG415 additionally contains single BamHI, BstEII and HincII sites and may also be used to clone PvuI-generated fragments; (5) the plasmids confer upon their host cells resistance to chloramphenicol, kanamycin and ampicillin, and every unique cloning site, except those of BamHI and BstEII, is located within one of these antibiotic-resistance genes.
Volume 16(1-3)
Pages 227-35
Published 1981-12-1
DOI 10.1016/0378-1119(81)90079-2
PII 0378-1119(81)90079-2
PMID 6282694
MeSH Anti-Bacterial Agents Bacteriophage lambda / genetics* Cloning, Molecular* Containment of Biohazards DNA Restriction Enzymes Drug Resistance, Microbial Escherichia coli / genetics* Genetic Vectors* Mutation Plasmids* Temperature
IF 2.984
Times Cited 163
WOS Category GENETICS & HEREDITY
Resource
Prokaryotes E. coli ME9034 pHSG415s pHSG422