1887

Abstract

Bifidobacteria are widely used as probiotics and have attracted increasing research interest worldwide. However, molecular techniques are still very scarce mainly due to the low efficiencies and strain-specific electroporation protocols that have been developed. Bacterial conjugation enables the transfer of genetic material among a relatively wide range of organisms and with virtually no size limitation. A conjugation protocol was developed based on the RP4 conjugative machinery in the strain WM3064(pBB109). Using this machinery, the newly constructed transmissible shuttle vector, pDOJHR-WD2, was successfully and consistently transferred into several strains representing four species at efficiencies which correlated with the to bifidobacteria ratios. Higher ratios were found to significantly improve transfer frequency per recipient, with almost 100 % transfer frequency occurring when the ratio was 10 : 1. The incompatible resident plasmid, pDOJH10S, in DJO10A was able to coexist, albeit at lower copy numbers, with the incoming vector pDOJHR-WD2 even though they possess the same . In some cases the copy number of this resident plasmid was too low to observe via gel electrophoresis, but it could be detected by Southern hybridization. Plasmid curing resulted in a strain, DJO10A-W3, that had lost both plasmids and this showed a one-log increase in conjugation efficiency due to the lack of plasmid incompatibility. In conclusion, this novel conjugative gene transfer protocol can be used for the introduction of genetic material (without size restriction) into species and is particularly useful for strains that are recalcitrant to electroporation.

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2013-02-01
2019-10-16
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References

  1. Al-Masaudi S. B., Russell A. D., Day M. J.. ( 1991;). Factors affecting conjugative transfer of plasmid pWG613, determining gentamicin resistance, in Staphylococcus aureus. . J Med Microbiol 34:, 103–107. [CrossRef][PubMed]
    [Google Scholar]
  2. Álvarez-Martín P., O’Connell-Motherway M., van Sinderen D., Mayo B.. ( 2007;). Functional analysis of the pBC1 replicon from Bifidobacterium catenulatum L48. . Appl Microbiol Biotechnol 76:, 1395–1402. [CrossRef][PubMed]
    [Google Scholar]
  3. Argnani A., Leer R. J., van Luijk N., Pouwels P. H.. ( 1996;). A convenient and reproducible method to genetically transform bacteria of the genus Bifidobacterium. . Microbiology 142:, 109–114. [CrossRef][PubMed]
    [Google Scholar]
  4. Bates S., Cashmore A. M., Wilkins B. M.. ( 1998;). IncP plasmids are unusually effective in mediating conjugation of Escherichia coli and Saccharomyces cerevisiae: involvement of the Tra2 mating system. . J Bacteriol 180:, 6538–6543.[PubMed]
    [Google Scholar]
  5. Blaesing F., Mühlenweg A., Vierling S., Ziegelin G., Pelzer S., Lanka E.. ( 2005;). Introduction of DNA into Actinomycetes by bacterial conjugation from E. coli–an evaluation of various transfer systems. . J Biotechnol 120:, 146–161. [CrossRef][PubMed]
    [Google Scholar]
  6. Bradley D. E.. ( 1980;). Morphological and serological relationships of conjugative pili. . Plasmid 4:, 155–169. [CrossRef][PubMed]
    [Google Scholar]
  7. Bradley D. E.. ( 1984;). Characteristics and function of thick and thin conjugative pili determined by transfer-derepressed plasmids of incompatibility groups I1, I2, I5, B, K and Z. . J Gen Microbiol 130:, 1489–1502.[PubMed]
    [Google Scholar]
  8. Bryan E. M., Bae T., Kleerebezem M., Dunny G. M.. ( 2000;). Improved vectors for nisin-controlled expression in Gram-positive bacteria. . Plasmid 44:, 183–190. [CrossRef][PubMed]
    [Google Scholar]
  9. Cascales E., Christie P. J.. ( 2004;). Definition of a bacterial type IV secretion pathway for a DNA substrate. . Science 304:, 1170–1173. [CrossRef][PubMed]
    [Google Scholar]
  10. Chiba K., Hoshino Y., Ishino K., Kogure T., Mikami Y., Uehara Y., Ishikawa J.. ( 2007;). Construction of a pair of practical NocardiaEscherichia coli shuttle vectors. . Jpn J Infect Dis 60:, 45–47.[PubMed]
    [Google Scholar]
  11. Crameri R., Davies J. E., Hütter R.. ( 1986;). Plasmid curing and generation of mutations induced with ethidium bromide in streptomycetes. . J Gen Microbiol 132:, 819–824.[PubMed]
    [Google Scholar]
  12. Datta N., Hedges R. W., Shaw E. J., Sykes R. B., Richmond M. H.. ( 1971;). Properties of an R factor from Pseudomonas aeruginosa. . J Bacteriol 108:, 1244–1249.[PubMed]
    [Google Scholar]
  13. Dehio C., Meyer M.. ( 1997;). Maintenance of broad-host-range incompatibility group P and group Q plasmids and transposition of Tn5 in Bartonella henselae following conjugal plasmid transfer from Escherichia coli. . J Bacteriol 179:, 538–540.[PubMed]
    [Google Scholar]
  14. Desomer J., Dhaese P., Van Montagu M.. ( 1988;). Conjugative transfer of cadmium resistance plasmids in Rhodococcus fascians strains. . J Bacteriol 170:, 2401–2405.[PubMed]
    [Google Scholar]
  15. Figurski D. H., Helinski D. R.. ( 1979;). Replication of an origin-containing derivative of plasmid RK2 dependent on a plasmid function provided in trans. . Proc Natl Acad Sci U S A 76:, 1648–1652. [CrossRef][PubMed]
    [Google Scholar]
  16. Fox G. E., Pechman K. R., Woese C. R.. ( 1977;). Comparative cataloging of 16S ribosomal ribonucleic acid: molecular approach to procaryotic systematics. . Int J Syst Bacteriol 27:, 44–57. [CrossRef]
    [Google Scholar]
  17. Gormley E. P., Davies J.. ( 1991;). Transfer of plasmid RSF1010 by conjugation from Escherichia coli to Streptomyces lividans and Mycobacterium smegmatis. . J Bacteriol 173:, 6705–6708.[PubMed]
    [Google Scholar]
  18. Hanahan D.. ( 1983;). Studies on transformation of Escherichia coli with plasmids. . J Mol Biol 166:, 557–580. [CrossRef][PubMed]
    [Google Scholar]
  19. Hirayama Y., Sakanaka M., Fukuma H., Murayama H., Kano Y., Fukiya S., Yokota A.. ( 2012;). Development of a double-crossover markerless gene deletion system in Bifidobacterium longum: functional analysis of the α-galactosidase gene for raffinose assimilation. . Appl Environ Microbiol 78:, 4984–4994. [CrossRef][PubMed]
    [Google Scholar]
  20. Isaacs F. J., Carr P. A., Wang H. H., Lajoie M. J., Sterling B., Kraal L., Tolonen A. C., Gianoulis T. A., Goodman D. B.. & other authors ( 2011;). Precise manipulation of chromosomes in vivo enables genome-wide codon replacement. . Science 333:, 348–353. [CrossRef][PubMed]
    [Google Scholar]
  21. Islam A.. ( 2006;). Iron reversible inhibition by bifidobacteria and microbial diversity of the human intestine. Masters thesis, University of Minnesota, Saint Paul, MN, USA.
  22. Kiatpapan P., Hashimoto Y., Nakamura H., Piao Y. Z., Ono H., Yamashita M., Murooka Y.. ( 2000;). Characterization of pRGO1, a plasmid from Propionibacterium acidipropionici, and its use for development of a host–vector system in propionibacteria. . Appl Environ Microbiol 66:, 4688–4695. [CrossRef][PubMed]
    [Google Scholar]
  23. Kim J. Y., Wang Y., Park M. S., Ji G. E.. ( 2010;). Improvement of transformation efficiency through in vitro methylation and SacII site mutation of plasmid vector in Bifidobacterium longum MG1. . J Microbiol Biotechnol 20:, 1022–1026. [CrossRef][PubMed]
    [Google Scholar]
  24. Kovach M. E., Elzer P. H., Hill D. S., Robertson G. T., Farris M. A., Roop R. M. II, Peterson K. M.. ( 1995;). Four new derivatives of the broad-host-range cloning vector pBBR1MCS, carrying different antibiotic-resistance cassettes. . Gene 166:, 175–176. [CrossRef][PubMed]
    [Google Scholar]
  25. Kullen M. J., Brady L. J., O’Sullivan D. J.. ( 1997;). Evaluation of using a short region of the recA gene for rapid and sensitive speciation of dominant bifidobacteria in the human large intestine. . FEMS Microbiol Lett 154:, 377–383. [CrossRef][PubMed]
    [Google Scholar]
  26. Lederberg J., Tatum E. L.. ( 1946;). Gene recombination in Escherichia coli. . Nature 158:, 558. [CrossRef][PubMed]
    [Google Scholar]
  27. Lee J. H., O’Sullivan D. J.. ( 2006;). Sequence analysis of two cryptic plasmids from Bifidobacterium longum DJO10A and construction of a shuttle cloning vector. . Appl Environ Microbiol 72:, 527–535. [CrossRef][PubMed]
    [Google Scholar]
  28. Lee J.-H., Karamychev V. N., Kozyavkin S. A., Mills D., Pavlov A. R., Pavlova N. V., Polouchine N. N., Richardson P. M., Shakhova V. V.. & other authors ( 2008;). Comparative genomic analysis of the gut bacterium Bifidobacterium longum reveals loci susceptible to deletion during pure culture growth. . BMC Genomics 9:, 247. [CrossRef][PubMed]
    [Google Scholar]
  29. Luzhetskyy A., Fedoryshyn M., Gromyko O., Ostash B., Rebets Y., Bechthold A., Fedorenko V.. ( 2006;). IncP plasmids are most effective in mediating conjugation between Escherichia coli and Streptomycetes. . Russ J Genet 42:, 476–481. [CrossRef]
    [Google Scholar]
  30. Matsumura H., Takeuchi A., Kano Y.. ( 1997;). Construction of Escherichia coli–Bifidobacterium longum shuttle vector transforming B. longum 105-A and 108-A. . Biosci Biotechnol Biochem 61:, 1211–1212. [CrossRef][PubMed]
    [Google Scholar]
  31. Mazodier P., Petter R., Thompson C.. ( 1989;). Intergeneric conjugation between Escherichia coli and Streptomyces species. . J Bacteriol 171:, 3583–3585.[PubMed]
    [Google Scholar]
  32. Mihajlovic S., Lang S., Sut M. V., Strohmaier H., Gruber C. J., Koraimann G., Cabezón E., Moncalián G., de la Cruz F., Zechner E. L.. ( 2009;). Plasmid r1 conjugative DNA processing is regulated at the coupling protein interface. . J Bacteriol 191:, 6877–6887. [CrossRef][PubMed]
    [Google Scholar]
  33. Moon G. S., Wegmann U., Gunning A. P., Gasson M. J., Narbad A.. ( 2009;). Isolation and characterization of a theta-type cryptic plasmid from Bifidobacterium longum FI10564. . J Microbiol Biotechnol 19:, 403–408. [CrossRef][PubMed]
    [Google Scholar]
  34. Nikodinovic J., Priestley N. D.. ( 2006;). A second generation snp-derived Escherichia coliStreptomyces shuttle expression vector that is generally transferable by conjugation. . Plasmid 56:, 223–227. [CrossRef][PubMed]
    [Google Scholar]
  35. Nordström K., Austin S. J.. ( 1989;). Mechanisms that contribute to the stable segregation of plasmids. . Annu Rev Genet 23:, 37–69. [CrossRef][PubMed]
    [Google Scholar]
  36. Novick R. P.. ( 1987;). Plasmid incompatibility. . Microbiol Rev 51:, 381–395.[PubMed]
    [Google Scholar]
  37. Novick R. P., Clowes R. C., Cohen S. N., Curtiss R. III, Datta N., Falkow S.. ( 1976;). Uniform nomenclature for bacterial plasmids: a proposal. . Bacteriol Rev 40:, 168–189.[PubMed]
    [Google Scholar]
  38. O’Connell Motherway M., O’Driscoll J., Fitzgerald G. F., Van Sinderen D.. ( 2009;). Overcoming the restriction barrier to plasmid transformation and targeted mutagenesis in Bifidobacterium breve UCC2003. . Microb Biotechnol 2:, 321–332. [CrossRef][PubMed]
    [Google Scholar]
  39. Palani N. P.. ( 2011;). Metabolic regulation and genetic tools for bacterial neutral lipid production. Masters thesis, University of Minnesota, Saint Paul, MN, USA.
  40. Pansegrau W., Balzer D., Kruft V., Lurz R., Lanka E.. ( 1990;). In vitro assembly of relaxosomes at the transfer origin of plasmid RP4. . Proc Natl Acad Sci U S A 87:, 6555–6559. [CrossRef][PubMed]
    [Google Scholar]
  41. Pansegrau W., Lanka E., Barth P. T., Figurski D. H., Guiney D. G., Haas D., Helinski D. R., Schwab H., Stanisich V. A., Thomas C. M.. ( 1994;). Complete nucleotide sequence of Birmingham IncP α plasmids. Compilation and comparative analysis. . J Mol Biol 239:, 623–663. [CrossRef][PubMed]
    [Google Scholar]
  42. Phornphisutthimas S., Sudtachat N., Bunyoo C., Chotewutmontri P., Panijpan B., Thamchaipenet A.. ( 2010;). Development of an intergeneric conjugal transfer system for rimocidin-producing Streptomyces rimosus. . Lett Appl Microbiol 50:, 530–536. [CrossRef][PubMed]
    [Google Scholar]
  43. Rauzier J., Moniz-Pereira J., Gicquel-Sanzey B.. ( 1988;). Complete nucleotide sequence of pAL5000, a plasmid from Mycobacterium fortuitum. . Gene 71:, 315–321. [CrossRef][PubMed]
    [Google Scholar]
  44. Rossi M., Brigidi P., Gonzalez Vara y Rodriguez A., Matteuzzi D.. ( 1996;). Characterization of the plasmid pMB1 from Bifidobacterium longum and its use for shuttle vector construction. . Res Microbiol 147:, 133–143. [CrossRef][PubMed]
    [Google Scholar]
  45. Rossi M., Brigidi P., Matteuzzi D.. ( 1998;). Improved cloning vectors for Bifidobacterium spp. . Lett Appl Microbiol 26:, 101–104. [CrossRef][PubMed]
    [Google Scholar]
  46. Schäfer A., Schwarzer A., Kalinowski J., Pühler A.. ( 1994;). Cloning and characterization of a DNA region encoding a stress-sensitive restriction system from Corynebacterium glutamicum ATCC 13032 and analysis of its role in intergeneric conjugation with Escherichia coli. . J Bacteriol 176:, 7309–7319.[PubMed]
    [Google Scholar]
  47. Schröder G., Lanka E.. ( 2005;). The mating pair formation system of conjugative plasmids–a versatile secretion machinery for transfer of proteins and DNA. . Plasmid 54:, 1–25. [CrossRef][PubMed]
    [Google Scholar]
  48. Shkoporov A. N., Efimov B. A., Khokhlova E. V., Steele J. L., Kafarskaia L. I., Smeianov V. V.. ( 2008a;). Characterization of plasmids from human infant Bifidobacterium strains: sequence analysis and construction of E. coli–Bifidobacterium shuttle vectors. . Plasmid 60:, 136–148. [CrossRef][PubMed]
    [Google Scholar]
  49. Smillie C., Garcillán-Barcia M. P., Francia M. V., Rocha E. P. C., de la Cruz F.. ( 2010;). Mobility of plasmids. . Microbiol Mol Biol Rev 74:, 434–452. [CrossRef][PubMed]
    [Google Scholar]
  50. Srivastava P., Singh P., Narayanan N., Deb J. K.. ( 2011;). Physiological and biochemical consequences of host–plasmid interaction–a case study with Corynebacterium renale, a multiple cryptic plasmid containing strain. . Plasmid 65:, 110–117. [CrossRef][PubMed]
    [Google Scholar]
  51. Szostková M., Horakova D.. ( 1998;). The effect of plasmid DNA sizes and other factors on electrotransformation of Escherichia coli JM109. . Bioelectrochem Bioenerg 47:, 319–323. [CrossRef]
    [Google Scholar]
  52. Thomas C. M., Smith C. A.. ( 1987;). Incompatibility group P plasmids: genetics, evolution, and use in genetic manipulation. . Annu Rev Microbiol 41:, 77–101. [CrossRef][PubMed]
    [Google Scholar]
  53. Tissier H.. ( 1899;). Le Bacterium coli et la reaction chromophile d'Escherich. . Crit Rev Soc Biol 51:, 943–945.
    [Google Scholar]
  54. Varsaki A., Moncalián G., del Pilar Garcillán-Barcia M., Drainas C., de la Cruz F.. ( 2009;). Analysis of ColE1 MbeC unveils an extended ribbon-helix-helix family of nicking accessory proteins. . J Bacteriol 191:, 1446–1455. [CrossRef][PubMed]
    [Google Scholar]
  55. Waters V. L.. ( 2001;). Conjugation between bacterial and mammalian cells. . Nat Genet 29:, 375–376. [CrossRef][PubMed]
    [Google Scholar]
  56. Yasui K., Kano Y., Tanaka K., Watanabe K., Shimizu-Kadota M., Yoshikawa H., Suzuki T.. ( 2009;). Improvement of bacterial transformation efficiency using plasmid artificial modification. . Nucleic Acids Res 37:, e3. [CrossRef][PubMed]
    [Google Scholar]
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