High-frequency electroporation and maintenance of pUC- and pBR-based cloning vectors in Pseudomonas stutzeri.
A number of Escherichia coli cloning vectors, based on ColE1-like replicons, were shown to be maintained in Pseudomonas stutzeri ATCC 17588. A restrictionless mutant of P. stutzeri was isolated, and this strain was used to develop an efficient electroporation system. With the E. coli cloning vector pHSG298, transformation frequencies of up to 2 x 10(7) transformants/micrograms DNA were achieved. This frequency is comparable to that obtained for CaCl2-mediated transformation of E. coli; thus, direct cloning of DNA into P. stutzeri is feasible. As will be discussed, this may prove useful for cloning DNA from high mol% G + C genera in cases in which E. coli is not a suitable heterologous cloning host.
Microbiology Department, University of Queensland, St. Lucia, Brisbane, Australia.
J M
JM
Pemberton
R J
RJ
Penfold
D003086:Bacteriocin Plasmids; D001431:Bacteriological Techniques; D002338:Carotenoids / Q000096:biosynthesis; D003001:Cloning, Molecular / Q000379:methods*; D004269:DNA, Bacterial / Q000235:genetics*; D004563:Electrochemistry; D005822:Genetic Vectors; D011549:Pseudomonas / Q000235:genetics*; D012093:Replicon; D014169:Transformation, Bacterial
D004269:DNA, Bacterial; D002338:Carotenoids