Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1981 Mar;145(3):1137–1143. doi: 10.1128/jb.145.3.1137-1143.1981

Molecular cloning of TOL genes xylB and xylE in Escherichia coli.

S Inouye, A Nakazawa, T Nakazawa
PMCID: PMC217113  PMID: 7009570

Abstract

The xylB and xylE genes in the TOL plasmid of Pseudomonas putida mt-2, which code for benzyl alcohol dehydrogenase and catechol 2,3-oxygenase, respectively, were cloned onto plasmid pBR322 in Escherichia coli for detailed mapping. The xylB gene was mapped in a 2.9-kilobase region within the BamHI BC fragment of pTN2, an in vivo RP4-TOL recombinant, whereas the xylE gene was mapped in a 1.8-kilobase region within the BamHI BD fragment. The directions of transcription of these genes were deduced from the expression of the cloned genes which had been ligated in orientations opposite pBR322 at its BamHI site within the tetracycline resistance gene. The xylB and xylE genes are inducible by a specific inducer of the TOL pathway genes in the RP4-TOL recombinant, whereas they are not inducible in the pBR322-TOL hybrids. The regulatory regions involved in expression of the xylB and xylE genes do not appear to be located in the vicinity of the structural genes. Catechol 2,3-oxygenase formed in E. coli carrying an xylE-containing plasmid is similar, or identical, to that formed in P. putida carrying the TOL plasmid.

Full text

PDF
1140

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bachmann B. J. Pedigrees of some mutant strains of Escherichia coli K-12. Bacteriol Rev. 1972 Dec;36(4):525–557. doi: 10.1128/br.36.4.525-557.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  3. Chang A. C., Cohen S. N. Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol. 1978 Jun;134(3):1141–1156. doi: 10.1128/jb.134.3.1141-1156.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Downing R., Broda P. A cleavage map of the TOL plasmid of Pseudomonas putida mt-2. Mol Gen Genet. 1979;177(1):189–191. doi: 10.1007/BF00267270. [DOI] [PubMed] [Google Scholar]
  5. Greene P. J., Heyneker H. L., Bolivar F., Rodriguez R. L., Betlach M. C., Covarrubias A. A., Backman K., Russel D. J., Tait R., Boyer H. W. A general method for the purification of restriction enzymes. Nucleic Acids Res. 1978 Jul;5(7):2373–2380. doi: 10.1093/nar/5.7.2373. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Miki T., Kimura M., Hiraga S., Nagata T., Yura T. Cloning and physical mapping of the dnaA region of the Escherichia coli chromosome. J Bacteriol. 1979 Dec;140(3):817–824. doi: 10.1128/jb.140.3.817-824.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Molholt B., Doskocil J. Increased transformation frequency in E. coli. Biochem Biophys Res Commun. 1978 May 30;82(2):477–483. doi: 10.1016/0006-291x(78)90899-9. [DOI] [PubMed] [Google Scholar]
  8. Murray N. E., Bruce S. A., Murray K. Molecular cloning of the DNA ligase gene from bacteriophage T4. II. Amplification and preparation of the gene product. J Mol Biol. 1979 Aug 15;132(3):493–505. doi: 10.1016/0022-2836(79)90271-7. [DOI] [PubMed] [Google Scholar]
  9. Nakazawa T., Hayashi E., Yokota T., Ebina Y., Nakazawa A. Isolation of TOL and RP4 recombinants by integrative suppression. J Bacteriol. 1978 Apr;134(1):270–277. doi: 10.1128/jb.134.1.270-277.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Nakazawa T., Inouye S., Nakazawa A. Physical and functional mapping of RP4-TOL plasmid recombinants: analysis of insertion and deletion mutants. J Bacteriol. 1980 Oct;144(1):222–231. doi: 10.1128/jb.144.1.222-231.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Nakazawa T., Yokota T. Benzoate metabolism in Pseudomonas putida(arvilla) mt-2: demonstration of two benzoate pathways. J Bacteriol. 1973 Jul;115(1):262–267. doi: 10.1128/jb.115.1.262-267.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Nozaki M. Oxygenases and dioxygenases. Top Curr Chem. 1979;78:145–186. doi: 10.1007/BFb0048193. [DOI] [PubMed] [Google Scholar]
  13. West R. W., Jr, Neve R. L., Rodriguez R. L. Construction and characterization of E. coli promoter-probe plasmid vectors. I. Cloning of promoter-containing DNA fragments. Gene. 1979 Nov;7(3-4):271–288. doi: 10.1016/0378-1119(79)90048-9. [DOI] [PubMed] [Google Scholar]
  14. Widera G., Gautier F., Lindenmaier W., Collins J. The expression of tetracycline resistance after insertion of foreign DNA fragments between the EcoRI and HindIII sites of the plasmid cloning vector pBR 322. Mol Gen Genet. 1978 Jul 25;163(3):301–305. doi: 10.1007/BF00271959. [DOI] [PubMed] [Google Scholar]
  15. Worsey M. J., Franklin F. C., Williams P. A. Regulation of the degradative pathway enzymes coded for by the TOL plasmid (pWWO) from Pseudomonas putida mt-2. J Bacteriol. 1978 Jun;134(3):757–764. doi: 10.1128/jb.134.3.757-764.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Worsey M. J., Williams P. A. Metabolism of toluene and xylenes by Pseudomonas (putida (arvilla) mt-2: evidence for a new function of the TOL plasmid. J Bacteriol. 1975 Oct;124(1):7–13. doi: 10.1128/jb.124.1.7-13.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES