Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1983 Sep;155(3):966–972. doi: 10.1128/jb.155.3.966-972.1983

A hybrid plasmid is a stable cloning vector for the cyanobacterium Anacystis nidulans R2.

S S Golden, L A Sherman
PMCID: PMC217787  PMID: 6309751

Abstract

Anacystis nidulans R2 is a highly transformable strain which is suitable as a recipient for molecular cloning in cyanobacteria. In an effort to produce an appropriate cloning vector, we constructed a hybrid plasmid molecule, pSG111, which contained pBR328 from Escherichia coli and the native pUH24 plasmid of A. nidulans. pSG111 replicated in and conferred ampicillin and chloramphenicol resistance to both hosts. It contained unique sites for the restriction enzymes EcoRI, SalI, SphI, and XhoI, which could be used for the insertion of exogenous DNA. To demonstrate that a molecule like pSG111 could serve as a shuttle vector for the cloning of A. nidulans genes, we constructed a hybrid plasmid, pRNA404, containing an A. nidulans rRNA operon. This recombinant molecule was genetically and structurally stable during passage through A. nidulans and E. coli. The stability of the hybrid plasmid and the inserted rRNA operon demonstrates the feasibility of cloning in A. nidulans with hybrid vectors, with the subsequent retrieval of cloned sequences.

Full text

PDF
971

Images in this article

968Image
on p.968
969Image
on p.969
970Image
on p.970

Selected References

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

  1. Blakesley R. W., Wells R. D. 'Single-stranded' DNA from phiX174 and M13 is cleaved by certain restriction endonucleases. Nature. 1975 Oct 2;257(5525):421–422. doi: 10.1038/257421a0. [DOI] [PubMed] [Google Scholar]
  2. Cohen S. N., Chang A. C., Hsu L. Nonchromosomal antibiotic resistance in bacteria: genetic transformation of Escherichia coli by R-factor DNA. Proc Natl Acad Sci U S A. 1972 Aug;69(8):2110–2114. doi: 10.1073/pnas.69.8.2110. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Goodman H. M., MacDonald R. J. Cloning of hormone genes from a mixture of cDNA molecules. Methods Enzymol. 1979;68:75–90. doi: 10.1016/0076-6879(79)68007-2. [DOI] [PubMed] [Google Scholar]
  4. Guikema J. A., Sherman L. A. Chlorophyll-protein organization of membranes from the cyanobacterium Anacystis nidulans. Arch Biochem Biophys. 1983 Jan;220(1):155–166. doi: 10.1016/0003-9861(83)90396-x. [DOI] [PubMed] [Google Scholar]
  5. Guikema J. A., Sherman L. A. Metronidazole and the isolation of temperature-sensitive photosynthetic mutants in cyanobacteria. J Bioenerg Biomembr. 1980 Aug;12(3-4):277–295. doi: 10.1007/BF00744689. [DOI] [PubMed] [Google Scholar]
  6. Holmes D. S., Quigley M. A rapid boiling method for the preparation of bacterial plasmids. Anal Biochem. 1981 Jun;114(1):193–197. doi: 10.1016/0003-2697(81)90473-5. [DOI] [PubMed] [Google Scholar]
  7. Kuhlemeier C. J., Borrias W. E., van den Hondel C. A., van Arkel G. A. Vectors for cloning in cyanobacteria: construction and characterization of two recombinant plasmids capable of transformation of Escherichia coli K12 and Anacystis nidulans R2. Mol Gen Genet. 1981;184(2):249–254. doi: 10.1007/BF00272912. [DOI] [PubMed] [Google Scholar]
  8. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  9. Sherman L. A., van de Putte P. Construction of a hybrid plasmid capable of replication in the bacterium Escherichia coli and the cyanobacterium Anacystis nidulans. J Bacteriol. 1982 Apr;150(1):410–413. doi: 10.1128/jb.150.1.410-413.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Shestakov S. V., Khyen N. T. Evidence for genetic transformation in blue-green alga Anacystis nidulans. Mol Gen Genet. 1970;107(4):372–375. doi: 10.1007/BF00441199. [DOI] [PubMed] [Google Scholar]
  11. Soberon X., Covarrubias L., Bolivar F. Construction and characterization of new cloning vehicles. IV. Deletion derivatives of pBR322 and pBR325. Gene. 1980 May;9(3-4):287–305. doi: 10.1016/0378-1119(90)90328-o. [DOI] [PubMed] [Google Scholar]
  12. 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]
  13. Tandeau de Marsac N., Borrias W. E., Kuhlemeier C. J., Castets A. M., van Arkel G. A., van den Hondel C. A. A new approach for molecular cloning in cyanobacteria: cloning of an Anacystis nidulans met gene using a Tn901-induced mutant. Gene. 1982 Nov;20(1):111–119. doi: 10.1016/0378-1119(82)90092-0. [DOI] [PubMed] [Google Scholar]
  14. van den Hondel C. A., Keegstra W., Borrias W. E., van Arkel G. A. Homology of plasmids in strains of unicellular Cyanobacteria. Plasmid. 1979 Jul;2(3):323–333. doi: 10.1016/0147-619x(79)90016-7. [DOI] [PubMed] [Google Scholar]
  15. van den Hondel C. A., Verbeek S., van der Ende A., Weisbeek P. J., Borrias W. E., van Arkel G. A. Introduction of transposon Tn901 into a plasmid of Anacystis nidulans: preparation for cloning in cyanobacteria. Proc Natl Acad Sci U S A. 1980 Mar;77(3):1570–1574. doi: 10.1073/pnas.77.3.1570. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES