1887

Abstract

The operon of the photosynthetic bacterium encodes reaction centre and B875 (LH1) antenna polypeptides of the photosynthetic apparatus. This region of the genome was used to establish the applicability of random transposon Tn5 mutagenesis in this bacterium. Four Tn5 insertions have been mapped and one of the mutants characterized using a variety of techniques in order to establish that the fluorescence properties and polypeptide composition were consistent with the absence of reaction centre polypeptides. Following the ‘rescue’ of the transposon along with flanking regions of the operon, re-introduction of this construction into wild-type yielded the original mutation. This demonstrates that following homologous recombination, localized mutagenesis can direct Tn5 into a predetermined region of the chromosome. Accordingly, genes borne on plasmid pSRC2 were mutagenized with Tn5 in , and the sites of insertion mapped physically. pSRC2 derivatives containing Tn5 were transferred to wild-type genes have been mapped by correlating the photosynthetic properties of resulting strains with the sites of Tn5 insertion into pSRC2.

Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-134-6-1481
1988-06-01
2021-05-17
Loading full text...

Full text loading...

/deliver/fulltext/micro/134/6/mic-134-6-1481.html?itemId=/content/journal/micro/10.1099/00221287-134-6-1481&mimeType=html&fmt=ahah

References

  1. Bauer C. E., Eleuterio M., Young D. A., Marrs B. L. 1987; Analysis of transcription through the Rhodobacter capsulatus puf operon using a translational fusion of puf M to the E. coli lac Z gene. In Progress in Photosynthesis Research 4 pp. 699–705 Biggins J. Edited by Dordrecht: Martinus Nijhoff;
    [Google Scholar]
  2. Biel A. J., Marrs B. L. 1983; Transcriptional regulation of several genes for bacteriochlorophyll biosynthesis in Rhodopseudomonas capsulata in response to oxygen. Journal of Bacteriology 156:686–694
    [Google Scholar]
  3. Boyer H. W., Roulland-Dussoix D. 1969; A complementation analysis of the restriction and modification of DNA in Escherichia coli. Journal of Molecular Biology 41:459–472
    [Google Scholar]
  4. De Brujin F. J., Lupski J. R. 1984; The use of transposon TnJ mutagenesis in the rapid generation of correlated physical and genetic maps of DNA segments cloned into multicopy plasmids - a review. Gene 27:131–149
    [Google Scholar]
  5. Chang A. C. Y., Cohen S. N. 1978; Construction and characterisation of amplifiable multicopy DNA cloning vehicles derived from the PI5A cryptic miniplasmid. Journal of Bacteriology 134:1141–1156
    [Google Scholar]
  6. Chory J., Kaplan S. 1982; The in vitro transcription-translation of DNA and RNA templates by extracts of Rhodopseudomonas sphaeroides. Journal of Biological Chemistry 257:15110–15121
    [Google Scholar]
  7. Donohue T. J., Mcewan A. G., Kaplan S. 1986; Cloning and expression of the Rhodobacter sphaeroides reaction centre H gene. Journal of Bacteriology 168:962–972
    [Google Scholar]
  8. Fornari C. S., Kaplan S. 1982; Genetic transformation of Rhodopseudomonas sphaeroides by plasmid DNA. Journal of Bacteriology 152:89–97
    [Google Scholar]
  9. Hanahan D. 1983; Studies on transformation of Escherichia coli with plasmids. Journal of Molecular Biology 166:557–580
    [Google Scholar]
  10. Hunter C. N., Turner G. 1988; Transfer of genes coding for apoproteins of reaction centre and lightharvesting LH1 complexes to Rhodobacter sphaeroides. Journal of General Microbiology 134:1471–1480
    [Google Scholar]
  11. Hunter C. N., Ashby M. K., Coomber S. A. 1987; The effect of oxygen on the levels of mRNA coding for reaction centre and light harvesting polypeptides of Rhodobacter sphaeroides. Biochemical Journal 247:489–492
    [Google Scholar]
  12. Jorgensen R. A., Rothstein S. J., Reznikoff W. S. 1979; A restriction enzyme cleavage map of Tn5 and location of a region encoding neomycin resistance. Molecular and General Genetics 177:65–72
    [Google Scholar]
  13. Kaufmann N., Hudig H., Drews G. 1984; Transposon Tn5 mutagenesis of genes for the photosynthetic apparatus in Rhodopseudomonas capsulata. Molecular and General Genetics 198:153–158
    [Google Scholar]
  14. Kramer H. J. M., Amesz J. 1982; Anisotropy of the emission and absorption bands of spinach chloroplasts measured by fluorescence polarization and polarized excitation spectra at low temperature. Biochimica et biophysica acta 682:201–207
    [Google Scholar]
  15. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature; London: 227:680–685
    [Google Scholar]
  16. Leong S. A., Ditta G. S., Helinski D. R. 1982; Heme biosynthesis in Rhizobium. Identification of a cloned gene coding for δ-aminolevulinic acid synthetase from Rhizobium meliloti. Journal of Biological Chemistry 257:8724–8730
    [Google Scholar]
  17. Nano F. E., Shepherd W. D., Watkins M. M., Kuhl S. A., Kaplan S. 1984; Broad-host-range plasmid vector for the in vitro construction of transcriptional/translational lac fusions. Gene 34:219–226
    [Google Scholar]
  18. Pemberton J. M., Bowen A. R. 1981; High-frequency chromosome transfer in Rhodopseudo- moms sphaeroides promoted by broad-host-range plasmid RP1 carrying mercury transposon Tn501. Journal of Bacteriology 147:110–117
    [Google Scholar]
  19. Simon R., Preifer U., Pühler A. 1983; A broad host range mobilization system for in vivo genetic engineering: transposon mutagenesis in Gram negative bacteria. Biotechmlogy 1:784–791
    [Google Scholar]
  20. Sistrom W. R. 1977; Transfer of chromosomal genes mediated by plasmid R68.45 in Rhodopseudomoms sphaeroides. Journal of Bacteriology 131:526–532
    [Google Scholar]
  21. Taylor D. P., Cohen S. N., Clark W. G., Marrs B. L. 1983; Alignment of genetic and restriction maps of the photosynthesis region of the Rhodopseudomoms capsulata chromosome by a conjugation-mediated marker rescue technique. Journal of Bacteriology 154:580–590
    [Google Scholar]
  22. Williams J. C., Steiner L. A., Ogden R. C., Simon M. I., Feher G. 1983; Primary structure of the M subunit of the reaction center from Rhodopseudomoms sphaeroides. Proceedings of the Natioml Academy of Sciences of the United States of America 80:6505–6509
    [Google Scholar]
  23. Williams J. C., Steiner L. A., Feher G., Simon M. I. 1984; Primary structure of the L subunit of the reaction center of Rhodopseudomoms sphaeroides. Proceedings of the Natioml Academy ofSciences of the United States of America 81:7303–7308
    [Google Scholar]
  24. Willison J., Ahombo G., Chabert J., Magnin J.-P., Vignais P. 1985; Genetic mapping of the Rhodopseudomoms capsulata chromosome shows non-clustering of genes involved in nitrogen fixation. Journal of General Microbiology 131:3001–3015
    [Google Scholar]
  25. Youvan D. C., Elder J. T., Sandlin D. E., Zsebo K., Alder D. P., Panopoulos N. J., Marrs B. L., Hearst J. E. 1982; R-prime site directed transposon Tn7 mutagenesis of the photosynthetic apparatus in Rhodopseudomoms capsulata. Journal of Molecular Biology 162:17–41
    [Google Scholar]
  26. Youvan D. C., Hearst J. E., Marrs B. L. 1983; Isolation and characterisation of enhanced fluorescence mutants of Rhodopseudomoms capsulata. Journal of Bacteriology 154:748–755
    [Google Scholar]
  27. Zhu Y. S., Kiley P. J., Donohue T. J., Kaplan S. 1986; Origin of the mRNA stoichiometry of the pufoperon in Rhodobacter sphaeroides. Journal of Biological Chemistry 261:10366–10374
    [Google Scholar]
  28. Zinchenko V. V., Babykin M. M., Shestakov S. V. 1984; Mobilization of non-conjugative plasmids into Rhodopseudomoms sphaeroides. Journal of General Microbiology 130:1587–1590
    [Google Scholar]
  29. Zsebo K. M., Hearst J. E. 1984; Genetic-physical mapping of a photosynthetic gene cluster from R. capsulata. Cell 37:937–947
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-134-6-1481
Loading
/content/journal/micro/10.1099/00221287-134-6-1481
Loading

Data & Media loading...

Most cited this month Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error