Summary
The Escherichia coli mutator gene mutT, which causes A:T→C:G transversion, was cloned in pBR 322. mutT + plasmids carry a 0.9 kb PvuII DNA fragment derived from the E. coli chromosome. Specific labelling of plasmid-encoded proteins by the maxicell method revealed that mutT codes for a polypeptide of about 15,000 daltons. The protein was overproduced when the mutT gene was placed under the control of the lac regulatory region on a multicopy runaway plasmid. The nucleotide sequence of the mutT gene was determined by the dideoxy method.
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Abbreviations
- Ap:
-
ampicillin
- IPTG:
-
isopropyl-β-d-thiogalactopyranoside
- kb:
-
kilobase pair(s)
- kDa:
-
kilodalton(s)
- SDS:
-
sodium dodecyl sulphate
- Tc:
-
tetracycline
References
Bachmann BJ (1983) Linkage map of Escherichia coli K-12, Edition 7. Microbiol Rev 47:180–230
Barnes WM, Bevan M, Son PH (1983) Kilo-sequencing: Creation of an ordered nest of asymmetric deletion across a large target sequence carried on phage M13. Methods Enzymol 101:98
Benton WD, Davis RW (1977) Screening λgt recombinant clones by hybridization to single plaques in situ. Science 196:180–182
Cox EC (1970) Mutator gene action and the replication of bacteriophage λ DNA. J Mol Biol 50:129–135
Cox EC (1973) Mutator gene studies in Escherichia coli: The mutT gene. Genet Suppl 73:67–80
Cox EC (1976) Bacterial mutator genes and the control of spontaneous mutation. Annu Rev Genet 10:135–156
Cox EC, Yanofsky C (1967) Altered base ratios in the DNA of an Escherichia coli mutator strain. Proc Natl Acad Sci USA 58:1895–1902
Cox EC, Yanofsky C (1969) Mutator gene studies in Escherichia coli. J Bacteriol 100:390–397
Degnen GE, Cox EC (1974) Conditional mutator gene in Escherichia coli: Isolation, mapping and effector studies. J Bacteriol 117:477–487
Guest JR, Stephens PE (1980) Molecular cloning of the pyruvate dehydrogenase complex genes of Escherichia coli. J Gen Microbiol 121:277–292
Hawley DK, McClure WR (1983) Compilation and analysis of Escherichia coli promoter DNA sequences. Nucleic Acids Res 11:2237–2255
Horiuchi T, Maki H, Sekiguchi M (1978) A new conditional lethal mutator (dnaQ49) in Escherichia coli K12. Mol Gen Genet 163:277–283
Horiuchi T, Maki H, Maruyama M, Sekiguchi M (1981) Identification of the dnaQ gene product and location of the structural gene for RNase H of Escherichia coli by cloning of the genes. Proc Natl Acad Sci USA 78:3770–3774
Horiuchi T, Maki H, Sekiguchi M (1984) RNase H-defective mutants of Escherichia coli: A possible discriminatory role of RNase H in initiation of DNA replication. Mol Gen Genet 195:17–22
Hübscher U, Lutz H, Kornberg A (1980) Novel histone H2A-like protein of Escherichia coli. Proc Natl Acad Sci USA 77:5097–5101
Kornberg A (1980) DNA replication. W.H. Freemen and Company, San Francisco
Kumura K, Sekiguchi M (1984) Identification of the uvrD gene product of Escherichia coli as DNA helicase II and its induction by DNA-damaging agents. J Biol Chem 259:1560–1565
Langley D, Guest JR (1978) Biochemical genetics of the α-keto acid dehydrogenase complexes of Escherichia coli K12: Genetic characterization and regulatory properties of deletion mutants. J Gen Microbiol 106:103–117
Lu A, Clark S, Modrich P (1983) Methyl-directed repair of DNA base-pair mismatches in vitro. Proc Natl Acad Sci USA 80:4639–4643
Maniatis T, Fritsch EF, Sambrook J (1982) Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York
Marmur J (1961) A procedure for the isolation of deoxyribonucleic acid from micro-organisms. J Mol Biol 3:208–218
Nakabeppu Y, Mine Y, Sekiguchi M (1985) Regulation of expression of the cloned ada gene in Escherichia coli. Mutat Res 146:155–167
Oeda K, Horiuchi T, Sekiguchi M (1981) Molecular cloning of the uvrD gene of Escherichia coli that controls ultraviolet sensitivity and spontaneous mutation frequency. Mol Gen Genet 184:191–199
Oeda K, Horiuchi T, Sekiguchi M (1982) The uvrD gene of E. coli encodes a DNA-dependent ATPase. Nature 298:98–100
Oliver DB, Beckwith J (1981) E. coli mutant pleiotropically defective in the export of secreted proteins. Cell 25:765–772
Oliver DB, Beckwith J (1982) Identification of a new gene (secA) and gene product involved in the secretion of envelope proteins in Escherichia coli. J Bacteriol 150:686–691
Perucho M, Goldfarb M, Shimizu K, Lama C, Fogh J, Wigler M (1981) Human-tumor-derived cell lines contain common and different transforming genes. Cell 27:467–476
Sancar A, Hack AM, Rupp WD (1979) Simple method for identification of plasmid-coded proteins. J Bacteriol 137:692–693
Sanger F, Coulson AR, Barrell BG, Smith AJH, Roe BA (1980) Cloning in single-stranded bacteriophage as an aid to rapid DNA sequencing. J Mol Biol 143:161–178
Scheuermann RH, Echols H (1984) A separate editing exonuclease for DNA replication: The ε subunit of Escherichia coli DNA polymerase III holoenzyme. Proc Natl Acad Sci USA 81:7747–7751
Shimizu K, Goldfarb M, Perucho M, Wigler M (1983) Isolation and preliminary characterization of the transforming gene of a human neuroblastoma cell line. Proc Natl Acad Sci USA 80:383–387
Shimizu K, Nakatsu Y, Sekiguchi M, Hokamura K, Tanaka K, Terada M, Sugimura T (1985) Molecular cloning of an activated human oncogene, homologous to v-raf, from primary stomach cancer. Proc Natl Acad Sci USA 82:5641–5645
Siegel EC, Kamel F (1974) Reversion of frameshift mutations by mutator genes in Escherichia coli. J Bacteriol 117:994–1001
Treffers HP, Spinelli V, Belser NO (1954) A factor (or mutator gene) influencing mutation rates in Escherichia coli. Proc Natl Acad Sci USA 40:1064–1071
Yamazaki K, Nagata A, Kano Y, Imamoto F (1984) Isolation and characterization of nucleoid proteins from Escherichia coli. Mol Gen Genet 196:217–224
Yanofsky C, Cox EC, Horn V (1966) The unusual mutagenic specificity of an E. coli mutator gene. Proc Natl Acad Sci USA 55:274–281
Watson MD (1985) A family of dispersed repetitive DNA in prokaryotes. TIBS 10:3–4
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Communicated by M. Takanami
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Akiyama, M., Horiuchi, T. & Sekiguchi, M. Molecular cloning and nucleotide sequence of the mutT mutator of Escherichia coli that causes A:T to C:G transversion. Mol Gen Genet 206, 9–16 (1987). https://doi.org/10.1007/BF00326530
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DOI: https://doi.org/10.1007/BF00326530