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. 1991 Aug;173(15):4683–4691. doi: 10.1128/jb.173.15.4683-4691.1991

Sequence and transcriptional regulation of com101A, a locus required for genetic transformation in Haemophilus influenzae.

T G Larson 1, S H Goodgal 1
PMCID: PMC208145  PMID: 1856167

Abstract

A 2.8-kb EcoRI-BglII fragment cloned from the wild-type Haemophilus influenzae Rd chromosome is shown to increase the transformability of the Com-101 mutant through trans complementation. Deletion and sequence analyses indicate that the active region of the clone carries a 687-bp open reading frame. A 0.3-kb insertion in the corresponding EcoRI-BglII fragment of the Com-101 chromosome is shown to be a partial (331-bp) duplication of this open reading frame. The wild-type sequence produces a peptide of a size that is consistent with the sequence data when this sequence is expressed in Escherichia coli with a T7 promoter-based transcription vector. RNA hybridization analysis using a DNA probe derived from the open reading frame suggests that the sequence is transiently expressed during competence development. On the basis of these observations, it is proposed that the open reading frame corresponds to the com101A gene.

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Selected References

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

  1. Albano M., Breitling R., Dubnau D. A. Nucleotide sequence and genetic organization of the Bacillus subtilis comG operon. J Bacteriol. 1989 Oct;171(10):5386–5404. doi: 10.1128/jb.171.10.5386-5404.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Albano M., Hahn J., Dubnau D. Expression of competence genes in Bacillus subtilis. J Bacteriol. 1987 Jul;169(7):3110–3117. doi: 10.1128/jb.169.7.3110-3117.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Barcak G. J., Tomb J. F., Laufer C. S., Smith H. O. Two Haemophilus influenzae Rd genes that complement the recA-like mutation rec-1. J Bacteriol. 1989 May;171(5):2451–2457. doi: 10.1128/jb.171.5.2451-2457.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Biggin M. D., Gibson T. J., Hong G. F. Buffer gradient gels and 35S label as an aid to rapid DNA sequence determination. Proc Natl Acad Sci U S A. 1983 Jul;80(13):3963–3965. doi: 10.1073/pnas.80.13.3963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Caster J. H., Postel E. H., Goodgal S. H. Competence mutants: isolation of transformation deficient strains of Haemophilus influenzae. Nature. 1970 Aug 1;227(5257):515–517. doi: 10.1038/227515a0. [DOI] [PubMed] [Google Scholar]
  7. Herriott R. M., Meyer E. M., Vogt M. Defined nongrowth media for stage II development of competence in Haemophilus influenzae. J Bacteriol. 1970 Feb;101(2):517–524. doi: 10.1128/jb.101.2.517-524.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Kauc L., Mitchell M., Goodgal S. H. Size and physical map of the chromosome of Haemophilus influenzae. J Bacteriol. 1989 May;171(5):2474–2479. doi: 10.1128/jb.171.5.2474-2479.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Larson T. G., Roszczyk E., Goodgal S. H. Molecular cloning of two linked loci that increase the transformability of transformation-deficient mutants of Haemophilus influenzae. J Bacteriol. 1991 Aug;173(15):4675–4682. doi: 10.1128/jb.173.15.4675-4682.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Lis J. T. Fractionation of DNA fragments by polyethylene glycol induced precipitation. Methods Enzymol. 1980;65(1):347–353. doi: 10.1016/s0076-6879(80)65044-7. [DOI] [PubMed] [Google Scholar]
  11. Mead D. A., Szczesna-Skorupa E., Kemper B. Single-stranded DNA 'blue' T7 promoter plasmids: a versatile tandem promoter system for cloning and protein engineering. Protein Eng. 1986 Oct-Nov;1(1):67–74. doi: 10.1093/protein/1.1.67. [DOI] [PubMed] [Google Scholar]
  12. Moffatt B. A., Studier F. W. T7 lysozyme inhibits transcription by T7 RNA polymerase. Cell. 1987 Apr 24;49(2):221–227. doi: 10.1016/0092-8674(87)90563-0. [DOI] [PubMed] [Google Scholar]
  13. Mohan S., Aghion J., Guillen N., Dubnau D. Molecular cloning and characterization of comC, a late competence gene of Bacillus subtilis. J Bacteriol. 1989 Nov;171(11):6043–6051. doi: 10.1128/jb.171.11.6043-6051.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Nakano M. M., Zuber P. Cloning and characterization of srfB, a regulatory gene involved in surfactin production and competence in Bacillus subtilis. J Bacteriol. 1989 Oct;171(10):5347–5353. doi: 10.1128/jb.171.10.5347-5353.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Panyim S., Chalkley R. The molecular weights of vertebrate histones exploiting a modified sodium dodecyl sulfate electrophoretic method. J Biol Chem. 1971 Dec 25;246(24):7557–7560. [PubMed] [Google Scholar]
  16. Parkison C., Cheng S. Y. A convenient method to increase the number of readable bases in DNA sequencing. Biotechniques. 1989 Sep;7(8):828–829. [PubMed] [Google Scholar]
  17. Penn M. D., Thireos G., Greer H. Temporal analysis of general control of amino acid biosynthesis in Saccharomyces cerevisiae: role of positive regulatory genes in initiation and maintenance of mRNA derepression. Mol Cell Biol. 1984 Mar;4(3):520–528. doi: 10.1128/mcb.4.3.520. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Poncz M., Solowiejczyk D., Ballantine M., Schwartz E., Surrey S. "Nonrandom" DNA sequence analysis in bacteriophage M13 by the dideoxy chain-termination method. Proc Natl Acad Sci U S A. 1982 Jul;79(14):4298–4302. doi: 10.1073/pnas.79.14.4298. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Studier F. W., Moffatt B. A. Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol. 1986 May 5;189(1):113–130. doi: 10.1016/0022-2836(86)90385-2. [DOI] [PubMed] [Google Scholar]
  21. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
  22. Stuy J. H., Walter R. B. Effect of glycerol on plasmid transfer in genetically competent Haemophilus influenzae. Mol Gen Genet. 1986 May;203(2):296–299. doi: 10.1007/BF00333969. [DOI] [PubMed] [Google Scholar]
  23. Tabor S., Richardson C. C. DNA sequence analysis with a modified bacteriophage T7 DNA polymerase. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4767–4771. doi: 10.1073/pnas.84.14.4767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Tomb J. F., Barcak G. J., Chandler M. S., Redfield R. J., Smith H. O. Transposon mutagenesis, characterization, and cloning of transformation genes of Haemophilus influenzae Rd. J Bacteriol. 1989 Jul;171(7):3796–3802. doi: 10.1128/jb.171.7.3796-3802.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Wang Y. Double-stranded DNA sequencing with T7 polymerase. Biotechniques. 1988 Oct;6(9):843–845. [PubMed] [Google Scholar]
  26. Weinrauch Y., Guillen N., Dubnau D. A. Sequence and transcription mapping of Bacillus subtilis competence genes comB and comA, one of which is related to a family of bacterial regulatory determinants. J Bacteriol. 1989 Oct;171(10):5362–5375. doi: 10.1128/jb.171.10.5362-5375.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]
  28. Youngman P., Zuber P., Perkins J. B., Sandman K., Igo M., Losick R. New ways to study developmental genes in spore-forming bacteria. Science. 1985 Apr 19;228(4697):285–291. doi: 10.1126/science.228.4697.285. [DOI] [PubMed] [Google Scholar]

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