Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1976 Oct;128(1):463–472. doi: 10.1128/jb.128.1.463-472.1976

Molecular cloning of an Escherichia coli plasmid determinant than encodes for the production of heat-stable enterotoxin.

M So, H W Boyer, M Betlach, S Falkow
PMCID: PMC232874  PMID: 789348

Abstract

A conjugative plasmid, ESF0041 was isolated from an enterotoxigenic strain of Escherichia coli from calves. ESF0041 was found to be 65 x 10(6) daltons in mass of a member of the F incompatibility complex. Acquisition of ESF0041 by E. coli K-12 was invariably associated with the capacity to produce heat-stable (ST) enterotoxin. ESF0041 and pSC101 deoxyribonucleic acids were cleaved with EcoRI, and the fragments were ligated with polynucleotide ligase. Transformation of E. coli K-12 with the ligation mixture led to the isolation of an ST+ clone. Further analysis of the plasmid deoxyribonucleic acid from this clone showed that a structural gene(s) associated with ST biosynthesis had been isolated as a 5.7 x 10(6)-dalton ESF0041 fragment in pSC101. In turn, 5.7 x 10(6)-dalton fragment was ligated to a multicopy COLE1 derivative, RSF2124, so that toxin synthesis was amplified about threefold.

Full text

PDF

Images in this article

Selected References

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

  1. Berg P., Baltimore D., Boyer H. W., Cohen S. N., Davis R. W., Hogness D. S., Nathans D., Roblin R., Watson J. D., Weissman S. Letter: Potential biohazards of recombinant DNA molecules. Science. 1974 Jul 26;185(4148):303–303. [PubMed] [Google Scholar]
  2. Berg P., Baltimore D., Brenner S., Roblin R. O., Singer M. F. Summary statement of the Asilomar conference on recombinant DNA molecules. Proc Natl Acad Sci U S A. 1975 Jun;72(6):1981–1984. doi: 10.1073/pnas.72.6.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Chang A. C., Cohen S. N. Genome construction between bacterial species in vitro: replication and expression of Staphylococcus plasmid genes in Escherichia coli. Proc Natl Acad Sci U S A. 1974 Apr;71(4):1030–1034. doi: 10.1073/pnas.71.4.1030. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Clewell D. B., Helinski D. R. Supercoiled circular DNA-protein complex in Escherichia coli: purification and induced conversion to an opern circular DNA form. Proc Natl Acad Sci U S A. 1969 Apr;62(4):1159–1166. doi: 10.1073/pnas.62.4.1159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Crosa J. H., Luttropp L. K., Falkow S. Nature of R-factor replication in the presence of chloramphenicol. Proc Natl Acad Sci U S A. 1975 Feb;72(2):654–658. doi: 10.1073/pnas.72.2.654. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dean A. G., Ching Y. C., Williams R. G., Harden L. B. Test for Escherichia coli enterotoxin using infant mice: application in a study of diarrhea in children in Honolulu. J Infect Dis. 1972 Apr;125(4):407–411. doi: 10.1093/infdis/125.4.407. [DOI] [PubMed] [Google Scholar]
  7. Dugaiczyk A., Hedgpeth J., Boyer H. W., Goodman H. M. Physical identity of the SV40 deoxyribonucleic acid sequence recognized by the Eco RI restriction endonuclease and modification methylase. Biochemistry. 1974 Jan 29;13(3):503–512. doi: 10.1021/bi00700a016. [DOI] [PubMed] [Google Scholar]
  8. Evans D. G., Silver R. P., Evans D. J., Jr, Chase D. G., Gorbach S. L. Plasmid-controlled colonization factor associated with virulence in Esherichia coli enterotoxigenic for humans. Infect Immun. 1975 Sep;12(3):656–667. doi: 10.1128/iai.12.3.656-667.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Falkow S., Williams L. P., Jr, Seaman S. L., Rollins L. D. Increased survival in calves of Escherichia coli K-12 carrying an Ent plasmid. Infect Immun. 1976 Mar;13(3):1005–1007. doi: 10.1128/iai.13.3.1005-1007.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Glover D. M., White R. L., Finnegan D. J., Hogness D. S. Characterization of six cloned DNAs from Drosophila melanogaster, including one that contains the genes for rRNA. Cell. 1975 Jun;5(2):149–157. doi: 10.1016/0092-8674(75)90023-9. [DOI] [PubMed] [Google Scholar]
  11. Guerrant R. L., Brunton L. L., Schnaitman T. C., Rebhun L. I., Gilman A. G. Cyclic adenosine monophosphate and alteration of Chinese hamster ovary cell morphology: a rapid, sensitive in vitro assay for the enterotoxins of Vibrio cholerae and Escherichia coli. Infect Immun. 1974 Aug;10(2):320–327. doi: 10.1128/iai.10.2.320-327.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gyles C. L., Barnum D. A. A heat-labile enterotoxin from strains of Eschericha coli enteropathogenic for pigs. J Infect Dis. 1969 Oct;120(4):419–426. doi: 10.1093/infdis/120.4.419. [DOI] [PubMed] [Google Scholar]
  13. Gyles C. L. Relationships among heat-labile enterotoxins of Escherichia coli and Vibrio cholerae. J Infect Dis. 1974 Mar;129(3):277–283. doi: 10.1093/infdis/129.3.277. [DOI] [PubMed] [Google Scholar]
  14. Gyles C., So M., Falkow S. The enterotoxin plasmids of Escherichia coli. J Infect Dis. 1974 Jul;130(1):40–49. doi: 10.1093/infdis/130.1.40. [DOI] [PubMed] [Google Scholar]
  15. Hershfield V., Boyer H. W., Yanofsky C., Lovett M. A., Helinski D. R. Plasmid ColEl as a molecular vehicle for cloning and amplification of DNA. Proc Natl Acad Sci U S A. 1974 Sep;71(9):3455–3459. doi: 10.1073/pnas.71.9.3455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Jacks T. M., Wu B. J. Biochemical properties of Escherichia coli low-molecular-weight, heat-stable enterotoxin. Infect Immun. 1974 Feb;9(2):342–347. doi: 10.1128/iai.9.2.342-347.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kedes L. H., Chang A. C., Houseman D., Cohen S. N. Isolation of histone genes from unfractionated sea urchin DNA by subculture cloning in E. coli. Nature. 1975 Jun 12;255(5509):533–538. doi: 10.1038/255533a0. [DOI] [PubMed] [Google Scholar]
  18. Keusch G. T., Grady G. F., Mata L. J., McIver J. The pathogenesis of Shigella diarrhea. I. Enterotoxin production by Shigella dysenteriae I. J Clin Invest. 1972 May;51(5):1212–1218. doi: 10.1172/JCI106915. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Morrow J. F., Cohen S. N., Chang A. C., Boyer H. W., Goodman H. M., Helling R. B. Replication and transcription of eukaryotic DNA in Escherichia coli. Proc Natl Acad Sci U S A. 1974 May;71(5):1743–1747. doi: 10.1073/pnas.71.5.1743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Smith H. W., Linggood M. A. Further observations on Escherichia coli enterotoxins with particular regard to those produced by atypical piglet strains and by calf and lamb strains: the transmissible nature of these enterotoxins and of a K antigen possessed by calf and lamb strains. J Med Microbiol. 1972 May;5(2):243–250. doi: 10.1099/00222615-5-2-243. [DOI] [PubMed] [Google Scholar]
  21. Smith H. W., Linggood M. A. Observations on the pathogenic properties of the K88, Hly and Ent plasmids of Escherichia coli with particular reference to porcine diarrhoea. J Med Microbiol. 1971 Nov;4(4):467–485. doi: 10.1099/00222615-4-4-467. [DOI] [PubMed] [Google Scholar]
  22. Smith H. W., Linggood M. A. The transmissible nature of enterotoxin production in a human enteropathogenic strain of Escherichia coli. J Med Microbiol. 1971 Aug;4(3):301–305. doi: 10.1099/00222615-4-3-301. [DOI] [PubMed] [Google Scholar]
  23. Smith N. W., Sack R. B. Immunologic cross-reactions of enterotoxins from Escherichia coli and Vibrio cholerae. J Infect Dis. 1973 Feb;127(2):164–170. doi: 10.1093/infdis/127.2.164. [DOI] [PubMed] [Google Scholar]
  24. So M., Crosa J. H., Falkow S. Polynucleotide sequence relationships among Ent plasmids and the relationship between Ent and other plasmids. J Bacteriol. 1975 Jan;121(1):234–238. doi: 10.1128/jb.121.1.234-238.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. So M., Gill R., Falkow S. The generation of a ColE1-Apr cloning vehicle which allows detection of inserted DNA. Mol Gen Genet. 1975 Dec 30;142(3):239–249. doi: 10.1007/BF00425649. [DOI] [PubMed] [Google Scholar]

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

RESOURCES