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. 1978 Mar;133(3):1254–1262. doi: 10.1128/jb.133.3.1254-1262.1978

Physiological factors involved in the transformation of Mycobacterium smegmatis.

M V Norgard, T Imaeda
PMCID: PMC222159  PMID: 641008

Abstract

Transfer of streptomycin resistance and changes from methionine and leucine auxotrophy to prototrophy were achieved in Mycobacterium smegmatis by transformation. Recipient cells were more resistant to mitomycin C and methyl methlanesulfonate treatments than were wild-type cells. A high level of calcium ions was essential for transformation, especially during DNA adsorption, whereas the presence of magnesium ions and the exposure of recipient cells to mild doses of UV light enhanced recombination frequencies. Transformants were not isolated when recipient cell-DNA mixtures were first treated with deoxyribonuclease. Recipient cells at various stages of growth showed similar transformabilities. Transformation was successful only when recipient cells were incubated on rich agar medium after mixture with DNA. Exposure of recipient cells to Pronase before treatment with donor DNA did not affect transformation, suggesting the absence of a protein competence factor. Throughout the present experiments, cotransformation frequencies were very low and unselected-marker segregation patterns were independent, indicating that the methionine, leucine, and streptomycin markers are not closely linked in M. smegmatis.

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Selected References

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

  1. Baess I. Isolation and purification of deoxyribonucleic acid from mycobacteria. Acta Pathol Microbiol Scand B Microbiol Immunol. 1974 Dec;82(6):780–784. doi: 10.1111/j.1699-0463.1974.tb02375.x. [DOI] [PubMed] [Google Scholar]
  2. Baxter-Gabbard K. L. A simple method for the large-scale preparation of sucrose gradients. FEBS Lett. 1972 Jan 15;20(1):117–119. doi: 10.1016/0014-5793(72)80031-0. [DOI] [PubMed] [Google Scholar]
  3. Brown D. D., Weber C. S. Gene linkage by RNA-DNA hybridization. II. Arrangement of the redundant gene sequences for 28 s and 18 s ribosomal RNA. J Mol Biol. 1968 Jun 28;34(3):681–697. doi: 10.1016/0022-2836(68)90189-7. [DOI] [PubMed] [Google Scholar]
  4. Carmody J. M., Herriott R. M. Thymine and thymidine uptake by Haemophilus influenzae and the labeling of deoxyribonucleic acid. J Bacteriol. 1970 Feb;101(2):525–530. doi: 10.1128/jb.101.2.525-530.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cosloy S. D., Oishi M. The nature of the transformation process in Escherichia coli K12. Mol Gen Genet. 1973 Jul 31;124(1):1–10. doi: 10.1007/BF00267159. [DOI] [PubMed] [Google Scholar]
  6. Dubnau D., Davidoff-Abelson R., Scher B., Cirigliano C. Fate of transforming deoxyribonucleic acid after uptake by competent Bacillus subtilis: phenotypic characterization of radiation-sensitive recombination-deficient mutants. J Bacteriol. 1973 Apr;114(1):273–286. doi: 10.1128/jb.114.1.273-286.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Freifelder D. Molecular weights of coliphages and coliphage DNA. IV. Molecular weights of DNA from bacteriophages T4, T5 and T7 and the general problem of determination of M. J Mol Biol. 1970 Dec 28;54(3):567–577. doi: 10.1016/0022-2836(70)90127-0. [DOI] [PubMed] [Google Scholar]
  8. Gelbart S. M., Juhasz S. E. Genetic transfer in Mycobacterium phlei. J Gen Microbiol. 1970 Dec;64(2):253–254. doi: 10.1099/00221287-64-2-253. [DOI] [PubMed] [Google Scholar]
  9. Gelbart S. M., Juhasz S. E. Transduction in mycobacterium phlei. Antonie Van Leeuwenhoek. 1973;39(1):1–10. doi: 10.1007/BF02578836. [DOI] [PubMed] [Google Scholar]
  10. Grange J. M. The genetics of mycobacteria and mycobacteriophages - a review. Tubercle. 1975 Sep;56(3):227–238. doi: 10.1016/0041-3879(75)90056-2. [DOI] [PubMed] [Google Scholar]
  11. Hawley R. J., Imaeda T., Mann N. Isolation and characterization of nocardia-like variants of Mycobacterium smegmatis. Can J Microbiol. 1976 Oct;22(10):1480–1491. doi: 10.1139/m76-219. [DOI] [PubMed] [Google Scholar]
  12. 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]
  13. Hopwood D. A. Genetic analysis and genome structure in Streptomyces coelicolor. Bacteriol Rev. 1967 Dec;31(4):373–403. doi: 10.1128/br.31.4.373-403.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. JUHASZ S. E. Interspecific hybridization among mycobacteria. Nature. 1960 Jan 23;185:265–265. doi: 10.1038/185265a0. [DOI] [PubMed] [Google Scholar]
  15. Jacob A. E., Hobbs S. J. Conjugal transfer of plasmid-borne multiple antibiotic resistance in Streptococcus faecalis var. zymogenes. J Bacteriol. 1974 Feb;117(2):360–372. doi: 10.1128/jb.117.2.360-372.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Joenje H., Konings W. N., Venema G. Interactions between exogenous deoxyribonucleic acid and membrane vesicles isolated from competent and noncompetent Bacillus subtilis. J Bacteriol. 1975 Mar;121(3):771–776. doi: 10.1128/jb.121.3.771-776.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jones W. D., David H. L. Preliminary observations on the occurrence of a streptomycin R-factor in Mycobacterium smegmatis ATCC 607. Tubercle. 1972 Mar;53(1):35–42. doi: 10.1016/0041-3879(72)90044-x. [DOI] [PubMed] [Google Scholar]
  18. Jones W. D., Jr, Beam R. E., David H. L. Transduction of a streptomycin R-factor from Mycobacterium smegmatis to Mycobacterium tuberculosis H37Rv. Tubercle. 1974 Mar;55(1):73–80. doi: 10.1016/0041-3879(74)90068-3. [DOI] [PubMed] [Google Scholar]
  19. KATUNUMA N., NAKASATO H. A study of the mechanism of the development of streptomycin resistant organisms by addition of the desoxyribonucleic acid prepared from resistant bacilli. Kekkaku. 1954 Jan;29(1):19–22. [PubMed] [Google Scholar]
  20. Lacks S., Neuberger M. Membrane location of a deoxyribonuclease implicated in the genetic transformation of Diplococcus pneumoniae. J Bacteriol. 1975 Dec;124(3):1321–1329. doi: 10.1128/jb.124.3.1321-1329.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lang D., Mitani M. Simplified quantitative electron microscopy of biopolymers. Biopolymers. 1970;9(3):373–379. doi: 10.1002/bip.1970.360090310. [DOI] [PubMed] [Google Scholar]
  22. Lang D. Molecular weights of coliphages and coliphage DNA. 3. Contour length and molecular weight of DNA from bacteriophages T4, T5 and T7, and from bovine papilloma virus. J Mol Biol. 1970 Dec 28;54(3):557–565. doi: 10.1016/0022-2836(70)90126-9. [DOI] [PubMed] [Google Scholar]
  23. Lederberg E. M., Cohen S. N. Transformation of Salmonella typhimurium by plasmid deoxyribonucleic acid. J Bacteriol. 1974 Sep;119(3):1072–1074. doi: 10.1128/jb.119.3.1072-1074.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Levinthal M. Bacterial genetics excluding E. coli. Annu Rev Microbiol. 1974;28(0):219–230. doi: 10.1146/annurev.mi.28.100174.001251. [DOI] [PubMed] [Google Scholar]
  25. Mizuguchi Y. Effect of ultraviolet-sensitive mutants on gene inheritance in mycobacterial matings. J Bacteriol. 1974 Feb;117(2):914–916. doi: 10.1128/jb.117.2.914-916.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Mizuguchi Y. Segregation of unselected markers in mycobacterial recombinants. Jpn J Microbiol. 1972 Mar;16(2):77–81. doi: 10.1111/j.1348-0421.1972.tb00632.x. [DOI] [PubMed] [Google Scholar]
  27. Mizuguchi Y., Tokunaga T. Method for isolation of deoxyribonucleic acid from mycobacteria. J Bacteriol. 1970 Nov;104(2):1020–1021. doi: 10.1128/jb.104.2.1020-1021.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Mizuguchi Y., Tokunaga T. Recombination between Mycobacterium smegmatis strains Jucho and Lacticola. Jpn J Microbiol. 1971 Jul;15(4):359–366. doi: 10.1111/j.1348-0421.1971.tb00592.x. [DOI] [PubMed] [Google Scholar]
  29. Notani N. K., Setlow J. K. Mechanism of bacterial transformation and transfection. Prog Nucleic Acid Res Mol Biol. 1974;14(0):39–100. doi: 10.1016/s0079-6603(08)60205-6. [DOI] [PubMed] [Google Scholar]
  30. Page W. J., Sadoff H. L. Physiological factors affecting transformation of Azotobacter vinelandii. J Bacteriol. 1976 Mar;125(3):1080–1087. doi: 10.1128/jb.125.3.1080-1087.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Raj C. V., Ramakrishnan T. Transduction in Mycobacterium smegmatis. Nature. 1970 Oct 17;228(5268):280–281. doi: 10.1038/228280b0. [DOI] [PubMed] [Google Scholar]
  32. Rudin L., Sjöström J. E., Lindberg M., Philipson L. Factors affecting competence for transformation in Staphylococcus aureus. J Bacteriol. 1974 Apr;118(1):155–164. doi: 10.1128/jb.118.1.155-164.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Schupp T., Hutter R., Hopwood D. A. Genetic recombination in Nocardia mediterranei. J Bacteriol. 1975 Jan;121(1):128–136. doi: 10.1128/jb.121.1.128-136.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sellers M. I., Tokunaga T. Further studies of infectious DNA extracted from mycobacteriophages. J Exp Med. 1966 Feb 1;123(2):327–340. doi: 10.1084/jem.123.2.327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Suga K., Mizuguchi Y. Mapping of antibiotic resistance markers in Mycobacterium smegmatis. Jpn J Microbiol. 1974 Mar;18(2):139–147. doi: 10.1111/j.1348-0421.1974.tb00802.x. [DOI] [PubMed] [Google Scholar]
  36. TOKUNAGA T., SELLERS M. INFECTION OF MYCOBACTERIUM SMEGMATIS WITH D29 PHAGE DNA. J Exp Med. 1964 Jan 1;119:139–149. doi: 10.1084/jem.119.1.139. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Tokunaga T., Mizuguchi Y., Suga K. Genetic recombination in mycobacteria. J Bacteriol. 1973 Mar;113(3):1104–1111. doi: 10.1128/jb.113.3.1104-1111.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Tokunaga T., Mizuguchi Y. [Sexuality and the genetics in mycobacteria (author's transl)]. Kekkaku. 1973 Oct;48(10):481–493. [PubMed] [Google Scholar]
  39. Tokunaga T., Nakamura R. M. Infection of competent Mycobacterium smegmatis with deoxyribonucleic acid extracted from bacteriophage B1. J Virol. 1968 Feb;2(2):110–117. doi: 10.1128/jvi.2.2.110-117.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Tárnok I., Bönicke R. Problems of genetic transformation of mycobacteria by deoxyribonucleic acid (DNA). Bull Int Union Tuberc. 1970 Jun;43:210–213. [PubMed] [Google Scholar]
  41. Witkin E. M. Ultraviolet mutagenesis and inducible DNA repair in Escherichia coli. Bacteriol Rev. 1976 Dec;40(4):869–907. doi: 10.1128/br.40.4.869-907.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]

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