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. 1983 Sep;155(3):1078–1087. doi: 10.1128/jb.155.3.1078-1087.1983

Cloning and mapping of the manganese superoxide dismutase gene (sodA) of Escherichia coli K-12.

D Touati
PMCID: PMC217801  PMID: 6309739

Abstract

An Escherichia coli gene bank composed of large DNA fragments (about 40 kilobases) was constructed by using the small cosmid pHC79. From it, a clone was isolated for its ability to overproduce superoxide dismutase. The enzyme overproduced was manganese superoxide dismutase, as determined by electrophoresis and antibody precipitation. Maxicell analysis and two-dimensional O'Farrell polyacrylamide gel electrophoresis demonstrated that the structural gene, sodA, of manganese superoxide dismutase was cloned. Subcloning fragments from the original cosmid located the sodA gene within a 4.8-kilobase EcoRI-BamHI fragment. This fragment was inserted into a lambda phage which was deleted for the att region and consequently could only lysogenize by recombination between the cloned bacterial DNA insertion and the bacterial chromosome. Genetic mapping of the prophage in such lysogens indicated that the chromosomal sodA locus lies near 87 min on the E. coli map.

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Selected References

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

  1. Bachmann B. J., Low K. B. Linkage map of Escherichia coli K-12, edition 6. Microbiol Rev. 1980 Mar;44(1):1–56. doi: 10.1128/mr.44.1.1-56.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Beauchamp C., Fridovich I. Superoxide dismutase: improved assays and an assay applicable to acrylamide gels. Anal Biochem. 1971 Nov;44(1):276–287. doi: 10.1016/0003-2697(71)90370-8. [DOI] [PubMed] [Google Scholar]
  3. 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]
  4. Blattner F. R., Williams B. G., Blechl A. E., Denniston-Thompson K., Faber H. E., Furlong L., Grunwald D. J., Kiefer D. O., Moore D. D., Schumm J. W. Charon phages: safer derivatives of bacteriophage lambda for DNA cloning. Science. 1977 Apr 8;196(4286):161–169. doi: 10.1126/science.847462. [DOI] [PubMed] [Google Scholar]
  5. Bridgen J., Harris J. I., Northrop F. Evolutionary relationships in superoxide dismutase. FEBS Lett. 1975 Jan 1;49(3):392–395. doi: 10.1016/0014-5793(75)80793-9. [DOI] [PubMed] [Google Scholar]
  6. Cameron J. R., Philippsen P., Davis R. W. Analysis of chromosomal integration and deletions of yeast plasmids. Nucleic Acids Res. 1977;4(5):1429–1448. doi: 10.1093/nar/4.5.1429. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Castillo C. J., Hsiao C. L., Coon P., Black L. W. Identification and perperties of bacteriophage T4 capsid-formation gene products. J Mol Biol. 1977 Mar 5;110(3):585–601. doi: 10.1016/s0022-2836(77)80113-7. [DOI] [PubMed] [Google Scholar]
  8. Clarke L., Carbon J. A colony bank containing synthetic Col El hybrid plasmids representative of the entire E. coli genome. Cell. 1976 Sep;9(1):91–99. doi: 10.1016/0092-8674(76)90055-6. [DOI] [PubMed] [Google Scholar]
  9. Collins J. Escherichia coli plasmids packageable in vitro in lambda bacteriophage particles. Methods Enzymol. 1979;68:309–326. doi: 10.1016/0076-6879(79)68022-9. [DOI] [PubMed] [Google Scholar]
  10. Cox E. C. Mutator gene action and the replication of bacteriophage lambda DNA. J Mol Biol. 1970 May 28;50(1):129–135. doi: 10.1016/0022-2836(70)90109-9. [DOI] [PubMed] [Google Scholar]
  11. Dougherty H. W., Sadowski S. J., Baker E. E. A new iron-containing superoxide dismutase from Escherichia coli. J Biol Chem. 1978 Jul 25;253(14):5220–5223. [PubMed] [Google Scholar]
  12. Enquist L., Tiemeier D., Leder P., Weisberg R., Sternberg N. Safer derivatives of bacteriophage lambdagt-lambdaC for use in cloning of recombinant DNA molecules. Nature. 1976 Feb 19;259(5544):596–598. doi: 10.1038/259596a0. [DOI] [PubMed] [Google Scholar]
  13. Fletcher G., Irwin C. A., Henson J. M., Fillingim C., Malone M. M., Walker J. R. Identification of the Escherichia coli cell division gene sep and organization of the cell division-cell envelope genes in the sep-mur-ftsA-envA cluster as determined with specialized transducing lambda bacteriophages. J Bacteriol. 1978 Jan;133(1):91–100. doi: 10.1128/jb.133.1.91-100.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fridovich I. Superoxide dismutases. Adv Enzymol Relat Areas Mol Biol. 1974;41(0):35–97. doi: 10.1002/9780470122860.ch2. [DOI] [PubMed] [Google Scholar]
  15. Garrels J. I. Two dimensional gel electrophoresis and computer analysis of proteins synthesized by clonal cell lines. J Biol Chem. 1979 Aug 25;254(16):7961–7977. [PubMed] [Google Scholar]
  16. George J., Castellazzi M., Buttin G. Prophage induction and cell division in E. coli. III. Mutations sfiA and sfiB restore division in tif and lon strains and permit the expression of mutator properties of tif. Mol Gen Genet. 1975 Oct 22;140(4):309–332. [PubMed] [Google Scholar]
  17. Gregory E. M., Fridovich I. Induction of superoxide dismutase by molecular oxygen. J Bacteriol. 1973 May;114(2):543–548. doi: 10.1128/jb.114.2.543-548.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Harris-Warrick R. M., Elkana Y., Ehrlich S. D., Lederberg J. Electrophoretic separation of Bacillus subtilis genes. Proc Natl Acad Sci U S A. 1975 Jun;72(6):2207–2211. doi: 10.1073/pnas.72.6.2207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hassan H. M., Fridovich I. Enzymatic defenses against the toxicity of oxygen and of streptonigrin in Escherichia coli. J Bacteriol. 1977 Mar;129(3):1574–1583. doi: 10.1128/jb.129.3.1574-1583.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hassan H. M., Fridovich I. Regulation of the synthesis of superoxide dismutase in Escherichia coli. Induction by methyl viologen. J Biol Chem. 1977 Nov 10;252(21):7667–7672. [PubMed] [Google Scholar]
  21. Hohn B., Collins J. A small cosmid for efficient cloning of large DNA fragments. Gene. 1980 Nov;11(3-4):291–298. doi: 10.1016/0378-1119(80)90069-4. [DOI] [PubMed] [Google Scholar]
  22. Hohn B. In vitro packaging of lambda and cosmid DNA. Methods Enzymol. 1979;68:299–309. doi: 10.1016/0076-6879(79)68021-7. [DOI] [PubMed] [Google Scholar]
  23. Howard-Flanders P., Boyce R. P. DNA repair and genetic recombination: studies on mutants of Escherichia coli defective in these processes. Radiat Res. 1966;(Suppl):156+–156+. [PubMed] [Google Scholar]
  24. Keele B. B., Jr, McCord J. M., Fridovich I. Superoxide dismutase from escherichia coli B. A new manganese-containing enzyme. J Biol Chem. 1970 Nov 25;245(22):6176–6181. [PubMed] [Google Scholar]
  25. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  26. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  27. Low K. B. Escherichia coli K-12 F-prime factors, old and new. Bacteriol Rev. 1972 Dec;36(4):587–607. doi: 10.1128/br.36.4.587-607.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Mandel M., Higa A. Calcium-dependent bacteriophage DNA infection. J Mol Biol. 1970 Oct 14;53(1):159–162. doi: 10.1016/0022-2836(70)90051-3. [DOI] [PubMed] [Google Scholar]
  29. Maral J., Puget K., Michelson A. M. Comparative study of superoxide dismutase, catalase and glutathione peroxidase levels in erythrocytes of different animals. Biochem Biophys Res Commun. 1977 Aug 22;77(4):1525–1535. doi: 10.1016/s0006-291x(77)80151-4. [DOI] [PubMed] [Google Scholar]
  30. Moustafa Hassan H., Fridovich I. Regulation of superoxide dismutase synthesis in Escherichia coli: glucose effect. J Bacteriol. 1977 Nov;132(2):505–510. doi: 10.1128/jb.132.2.505-510.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
  32. OUCHTERLONY O. Diffusion-in-gel methods for immunological analysis. II. Prog Allergy. 1962;6:30–154. doi: 10.1159/000313795. [DOI] [PubMed] [Google Scholar]
  33. Pahel G., Bloom F. R., Tyler B. Deletion mapping of the polA-metB region of the Escherichia coli chromosome. J Bacteriol. 1979 May;138(2):653–656. doi: 10.1128/jb.138.2.653-656.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Plumbridge J. A., Howe J. G., Springer M., Touati-Schwartz D., Hershey J. W., Grunberg-Manago M. Cloning and mapping of a gene for translational initiation factor IF2 in Escherichia coli. Proc Natl Acad Sci U S A. 1982 Aug;79(16):5033–5037. doi: 10.1073/pnas.79.16.5033. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Rupp W. D., Wilde C. E., 3rd, Reno D. L., Howard-Flanders P. Exchanges between DNA strands in ultraviolet-irradiated Escherichia coli. J Mol Biol. 1971 Oct 14;61(1):25–44. doi: 10.1016/0022-2836(71)90204-x. [DOI] [PubMed] [Google Scholar]
  36. Sancar A., Hack A. M., Rupp W. D. Simple method for identification of plasmid-coded proteins. J Bacteriol. 1979 Jan;137(1):692–693. doi: 10.1128/jb.137.1.692-693.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Shimada K., Weisberg R. A., Gottesman M. E. Prophage lambda at unusual chromosomal locations. I. Location of the secondary attachment sites and the properties of the lysogens. J Mol Biol. 1972 Feb 14;63(3):483–503. doi: 10.1016/0022-2836(72)90443-3. [DOI] [PubMed] [Google Scholar]
  38. Steinman H. M., Hill R. L. Sequence homologies among bacterial and mitochondrial superoxide dismutases. Proc Natl Acad Sci U S A. 1973 Dec;70(12):3725–3729. doi: 10.1073/pnas.70.12.3725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Studier F. W. Analysis of bacteriophage T7 early RNAs and proteins on slab gels. J Mol Biol. 1973 Sep 15;79(2):237–248. doi: 10.1016/0022-2836(73)90003-x. [DOI] [PubMed] [Google Scholar]
  40. Thompson R., Hughes S. G., Broda P. Plasmid identification using specific endonucleases. Mol Gen Genet. 1974;133(2):141–149. doi: 10.1007/BF00264835. [DOI] [PubMed] [Google Scholar]
  41. Wessler S. R., Calvo J. M. Control of leu operon expression in Escherichia coli by a transcription attenuation mechanism. J Mol Biol. 1981 Jul 15;149(4):579–597. doi: 10.1016/0022-2836(81)90348-x. [DOI] [PubMed] [Google Scholar]
  42. Wu T. T. A model for three-point analysis of random general transduction. Genetics. 1966 Aug;54(2):405–410. doi: 10.1093/genetics/54.2.405. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Yamamoto K. R., Alberts B. M., Benzinger R., Lawhorne L., Treiber G. Rapid bacteriophage sedimentation in the presence of polyethylene glycol and its application to large-scale virus purification. Virology. 1970 Mar;40(3):734–744. doi: 10.1016/0042-6822(70)90218-7. [DOI] [PubMed] [Google Scholar]
  44. Yost F. J., Jr, Fridovich I. An iron-containing superoxide dismutase from Escherichia coli. J Biol Chem. 1973 Jul 25;248(14):4905–4908. [PubMed] [Google Scholar]

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