Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1982 Oct;152(1):215–222. doi: 10.1128/jb.152.1.215-222.1982

Helper plasmid cloning in Streptococcus sanguis: cloning of a tetracycline resistance determinant from the Streptococcus mutans chromosome.

J A Tobian, F L Macrina
PMCID: PMC221394  PMID: 6288658

Abstract

A model system for testing the helper plasmid cloning system of Gryczan et al. (Mol. Gen. Genet. 177:459-467, 1980) was devised for the Streptococcus sanguis (Challis) host-vector system. In this system, linearized pVA736 plasmid efficiently transformed an S. sanguis (Challis) host containing a homologous plasmid, pVA380-1, but did not transform a plasmidless host or a host containing a nonhomologous plasmid, pVA380. In addition, whereas monomeric circular pVA736 transformed a plasmidless host with two-hit kinetics, it transformed a pVA380-1-containing host with one-hit kinetics. This helper plasmid cloning system was used to isolate two HindIII fragments (5.0 megadaltons [Mdal] and 1.9 Mdal in size) from the chromosome of Streptococcus mutans V825 which conferred high-level tetracycline resistance. One tetracycline-resistant clone was examined and found to contain three plasmids which were sized and designated pVA868 (9.0 Mdal), pVA869 (9.5 Mdal), and pVA870 (9.8 Mdal). Results of Southern blot hybridization and restriction endonuclease digestion confirmed that all three chimeras were composed of two HindIII fragments of the S. mutans V825 chromosome, as well as a large portion, varying in size for each chimera, of the 2.8 Mdal cloning vector, pVA380-1. Incompatibility observed between pVA380-1 and each of the chimeras indicated that replication of the chimeras was governed by the pVA380-1 replicative origin. Southern blotting experiments revealed that the chimeras hybridized to Tn916, providing the first evidence that transposon-related genes of enteric streptococcal origin are disseminated among oral streptococci.

Full text

PDF
215

Images in this article

219Image
on p.219
219Image
on p.219

Selected References

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

  1. Behnke D., Ferretti J. J. Molecular cloning of an erythromycin resistance determinant in streptococci. J Bacteriol. 1980 Nov;144(2):806–813. doi: 10.1128/jb.144.2.806-813.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Behnke D., Gilmore M. S. Location of antibiotic resistance determinants, copy control, and replication functions on the double-selective streptococcal cloning vector pGB301. Mol Gen Genet. 1981;184(1):115–120. doi: 10.1007/BF00271206. [DOI] [PubMed] [Google Scholar]
  3. Behnke D. Plasmid transformation of Streptococcus sanguis (Challis) occurs by circular and linear molecules. Mol Gen Genet. 1981;182(3):490–497. doi: 10.1007/BF00293940. [DOI] [PubMed] [Google Scholar]
  4. Franke A. E., Clewell D. B. Evidence for a chromosome-borne resistance transposon (Tn916) in Streptococcus faecalis that is capable of "conjugal" transfer in the absence of a conjugative plasmid. J Bacteriol. 1981 Jan;145(1):494–502. doi: 10.1128/jb.145.1.494-502.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Gryczan T., Contente S., Dubnau D. Molecular cloning of heterologous chromosomal DNA by recombination between a plasmid vector and a homologous resident plasmid in Bacillus subtilis. Mol Gen Genet. 1980 Feb;177(3):459–467. doi: 10.1007/BF00271485. [DOI] [PubMed] [Google Scholar]
  6. Hansen J. B., Olsen R. H. Isolation of large bacterial plasmids and characterization of the P2 incompatibility group plasmids pMG1 and pMG5. J Bacteriol. 1978 Jul;135(1):227–238. doi: 10.1128/jb.135.1.227-238.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Lacks S. Uptake of circular deoxyribonucleic acid and mechanism of deoxyribonucleic acid transport in genetic transformation of Streptococcus pneumoniae. J Bacteriol. 1979 May;138(2):404–409. doi: 10.1128/jb.138.2.404-409.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Lawson J. W., Gooder H. Growth and development of competence in the group H streptococci. J Bacteriol. 1970 Jun;102(3):820–825. doi: 10.1128/jb.102.3.820-825.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Littleton N. W., Kakehashi S., Fitzgerald R. J. Recovery of specific "caries-inducing" streptococci from carious lesions in the teeth of children. Arch Oral Biol. 1970 May;15(5):461–463. doi: 10.1016/0003-9969(70)90073-7. [DOI] [PubMed] [Google Scholar]
  10. Loesche W. J., Rowan J., Straffon L. H., Loos P. J. Association of Streptococcus mutants with human dental decay. Infect Immun. 1975 Jun;11(6):1252–1260. doi: 10.1128/iai.11.6.1252-1260.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Macrina F. L., Jones K. R., Welch R. A. Transformation of Streptococcus sanguis with monomeric pVA736 plasmid deoxyribonucleic acid. J Bacteriol. 1981 May;146(2):826–830. doi: 10.1128/jb.146.2.826-830.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Macrina F. L., Jones K. R., Wood P. H. Chimeric streptococcal plasmids and their use as molecular cloning vehicles in Streptococcus sanguis (Challis). J Bacteriol. 1980 Sep;143(3):1425–1435. doi: 10.1128/jb.143.3.1425-1435.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Macrina F. L., Kopecko D. J., Jones K. R., Ayers D. J., McCowen S. M. A multiple plasmid-containing Escherichia coli strain: convenient source of size reference plasmid molecules. Plasmid. 1978 Jun;1(3):417–420. doi: 10.1016/0147-619x(78)90056-2. [DOI] [PubMed] [Google Scholar]
  14. Macrina F. L., Wood P. H., Jones K. R. Genetic transformation of Streptococcus sanguis (Challis) with cryptic plasmids from Streptococcus ferus. Infect Immun. 1980 Jun;28(3):692–699. doi: 10.1128/iai.28.3.692-699.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Macrina F. L., Wood P. H., Jones K. R. Simple method for demonstrating small plasmid deoxyribonucleic acid molecules in oral streptococci. Appl Environ Microbiol. 1980 May;39(5):1070–1073. doi: 10.1128/aem.39.5.1070-1073.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Malke H., Burman L. G., Holm S. E. Molecular cloning in streptococci: physical mapping of the vehicle plasmid pSM10 and demonstration of intergroup DNA transfer. Mol Gen Genet. 1981;181(2):259–267. doi: 10.1007/BF00268435. [DOI] [PubMed] [Google Scholar]
  17. Meyers J. A., Sanchez D., Elwell L. P., Falkow S. Simple agarose gel electrophoretic method for the identification and characterization of plasmid deoxyribonucleic acid. J Bacteriol. 1976 Sep;127(3):1529–1537. doi: 10.1128/jb.127.3.1529-1537.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Saunders C. W., Guild W. R. Monomer plasmid DNA transforms Streptococcus pneumoniae. Mol Gen Genet. 1981;181(1):57–62. doi: 10.1007/BF00339005. [DOI] [PubMed] [Google Scholar]
  19. Saunders C. W., Guild W. R. Pathway of plasmid transformation in Pneumococcus: open circular and linear molecules are active. J Bacteriol. 1981 May;146(2):517–526. doi: 10.1128/jb.146.2.517-526.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Saunders C. W., Guild W. R. Properties and transforming activities of two plasmids in Streptococcus pneumoniae. Mol Gen Genet. 1980;180(3):573–578. doi: 10.1007/BF00268062. [DOI] [PubMed] [Google Scholar]
  21. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  22. Strike P., Humphreys G. O., Roberts R. J. Nature of transforming deoxyribonucleic acid in calcium-treated Escherichia coli. J Bacteriol. 1979 Jun;138(3):1033–1035. doi: 10.1128/jb.138.3.1033-1035.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Tanaka T., Weisblum B., Schnös M., Inman R. Construction and characterization of a chimeric plasmid composed of DNA Pfrom Escherichia coli and Drosophila melanogaster. Biochemistry. 1975 May 20;14(10):2064–2072. doi: 10.1021/bi00681a005. [DOI] [PubMed] [Google Scholar]

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

RESOURCES