Skip to main content
NCBI home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1986 Mar 11;14(5):2365–2380. doi: 10.1093/nar/14.5.2365

Mutations induced by DNA polymerase alpha upon in vitro replication of M13mp8(+) DNA.

B Reckmann, G Krauss
PMCID: PMC339663  PMID: 3960723

Abstract

The forward mutation of the lacZ part of the bacteriophage M13mp8 has been used to study the fidelity of the 9S DNA polymerase alpha from calf thymus during in vitro replication of single-stranded DNA. Errors leading to a loss of alpha-complementation were identified by DNA sequencing. The overall mutation rate of the lacZ target sequence was in the range of 1:300-1:1000 which is more than one order of magnitude higher than the spontaneous mutation rate. In a mutL host the mutation rate was nearly threefold higher as compared to the wildtype host. Base substitutions comprise 86% of the errors whereas base deletions amount to 12%. The addition of a base was detected only in one mutant out of 71 sequenced ones. The frameshift mutations occurred predominantly in runs of the same base. The frequencies of individual base substitution are in the order of 2 X 10(-4)-4 X 10(-4) for most of the mismatches. Mutations involving dCTP:T and dGTP:T mismatches are observed with a lower frequency, those involving dTTP:C mismatches with a higher frequency.

Full text

PDF
2365

Selected References

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

  1. Fersht A. R., Knill-Jones J. W. Fidelity of replication of bacteriophage phi X174 DNA in vitro and in vivo. J Mol Biol. 1983 Apr 25;165(4):633–654. doi: 10.1016/s0022-2836(83)80271-x. [DOI] [PubMed] [Google Scholar]
  2. Fersht A. R., Shi J. P., Tsui W. C. Kinetics of base misinsertion by DNA polymerase I of Escherichia coli. J Mol Biol. 1983 Apr 25;165(4):655–667. doi: 10.1016/s0022-2836(83)80272-1. [DOI] [PubMed] [Google Scholar]
  3. Frank R., Heikens W., Heisterberg-Moutsis G., Blöcker H. A new general approach for the simultaneous chemical synthesis of large numbers of oligonucleotides: segmental solid supports. Nucleic Acids Res. 1983 Jul 11;11(13):4365–4377. doi: 10.1093/nar/11.13.4365. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Glickman B. W., Radman M. Escherichia coli mutator mutants deficient in methylation-instructed DNA mismatch correction. Proc Natl Acad Sci U S A. 1980 Feb;77(2):1063–1067. doi: 10.1073/pnas.77.2.1063. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Godson G. N., Vapnek D. A simple method of preparing large amounts of phiX174 RF 1 supercoiled DNA. Biochim Biophys Acta. 1973 Apr 11;299(4):516–520. doi: 10.1016/0005-2787(73)90223-2. [DOI] [PubMed] [Google Scholar]
  6. Grosse F., Krauss G., Knill-Jones J. W., Fersht A. R. Accuracy of DNA polymerase-alpha in copying natural DNA. EMBO J. 1983;2(9):1515–1519. doi: 10.1002/j.1460-2075.1983.tb01616.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Grosse F., Krauss G. Purification of a 9S DNA polymerase alpha species from calf thymus. Biochemistry. 1981 Sep 15;20(19):5470–5475. doi: 10.1021/bi00522a019. [DOI] [PubMed] [Google Scholar]
  8. Grosse F., Krauss G. Replication of M13mp7 single-stranded DNA in vitro by the 9-S DNA polymerase alpha from calf thymus. Eur J Biochem. 1984 May 15;141(1):109–114. doi: 10.1111/j.1432-1033.1984.tb08164.x. [DOI] [PubMed] [Google Scholar]
  9. Grosse F., Krauss G. The primase activity of DNA polymerase alpha from calf thymus. J Biol Chem. 1985 Feb 10;260(3):1881–1888. [PubMed] [Google Scholar]
  10. Herman G. E., Modrich P. Escherichia coli dam methylase. Physical and catalytic properties of the homogeneous enzyme. J Biol Chem. 1982 Mar 10;257(5):2605–2612. [PubMed] [Google Scholar]
  11. Hibner U., Alberts B. M. Fidelity of DNA replication catalysed in vitro on a natural DNA template by the T4 bacteriophage multi-enzyme complex. Nature. 1980 May 29;285(5763):300–305. doi: 10.1038/285300a0. [DOI] [PubMed] [Google Scholar]
  12. Hill R. F. Location of genes controlling excision repair of UV damage and mutator activity in Escherichia coli WP2. Mutat Res. 1970 Mar;9(3):341–344. doi: 10.1016/0027-5107(70)90135-1. [DOI] [PubMed] [Google Scholar]
  13. Kramer B., Kramer W., Fritz H. J. Different base/base mismatches are corrected with different efficiencies by the methyl-directed DNA mismatch-repair system of E. coli. Cell. 1984 Oct;38(3):879–887. doi: 10.1016/0092-8674(84)90283-6. [DOI] [PubMed] [Google Scholar]
  14. Kramer W., Schughart K., Fritz H. J. Directed mutagenesis of DNA cloned in filamentous phage: influence of hemimethylated GATC sites on marker recovery from restriction fragments. Nucleic Acids Res. 1982 Oct 25;10(20):6475–6485. doi: 10.1093/nar/10.20.6475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kunkel T. A. Mutational specificity of depurination. Proc Natl Acad Sci U S A. 1984 Mar;81(5):1494–1498. doi: 10.1073/pnas.81.5.1494. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kunkel T. A. The mutational specificity of DNA polymerase-beta during in vitro DNA synthesis. Production of frameshift, base substitution, and deletion mutations. J Biol Chem. 1985 May 10;260(9):5787–5796. [PubMed] [Google Scholar]
  17. LeClerc J. E., Istock N. L., Saran B. R., Allen R., Jr Sequence analysis of ultraviolet-induced mutations in M13lacZ hybrid phage DNA. J Mol Biol. 1984 Dec 5;180(2):217–237. doi: 10.1016/s0022-2836(84)80001-7. [DOI] [PubMed] [Google Scholar]
  18. LeClerc J. E., Istock N. L. Specificity of UV mutagenesis in the lac promoter of M13lac hybrid phage DNA. Nature. 1982 Jun 17;297(5867):596–598. doi: 10.1038/297596a0. [DOI] [PubMed] [Google Scholar]
  19. Loeb L. A., Kunkel T. A. Fidelity of DNA synthesis. Annu Rev Biochem. 1982;51:429–457. doi: 10.1146/annurev.bi.51.070182.002241. [DOI] [PubMed] [Google Scholar]
  20. Lu A. L., Clark S., Modrich P. Methyl-directed repair of DNA base-pair mismatches in vitro. Proc Natl Acad Sci U S A. 1983 Aug;80(15):4639–4643. doi: 10.1073/pnas.80.15.4639. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Marinus M. G., Morris N. R. Pleiotropic effects of a DNA adenine methylation mutation (dam-3) in Escherichia coli K12. Mutat Res. 1975 Apr;28(1):15–26. doi: 10.1016/0027-5107(75)90309-7. [DOI] [PubMed] [Google Scholar]
  22. Messing J., Gronenborn B., Müller-Hill B., Hans Hopschneider P. Filamentous coliphage M13 as a cloning vehicle: insertion of a HindII fragment of the lac regulatory region in M13 replicative form in vitro. Proc Natl Acad Sci U S A. 1977 Sep;74(9):3642–3646. doi: 10.1073/pnas.74.9.3642. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Rienitz A., Grosse F., Blöcker H., Frank R., Krauss G. On the fidelity of DNA polymerase alpha: the influence of alpha-thio dNTPs, Mn2+ and mismatch repair. Nucleic Acids Res. 1985 Aug 12;13(15):5685–5695. doi: 10.1093/nar/13.15.5685. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. 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]
  25. Siegel E. C., Kamel F. Reversion of frameshift mutations by mutator genes in Escherichia coli. J Bacteriol. 1974 Mar;117(3):994–1001. doi: 10.1128/jb.117.3.994-1001.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Soltis D. A., Lehman I. R. recA protein promoted DNA strand exchange. J Biol Chem. 1983 May 25;258(10):6073–6077. [PubMed] [Google Scholar]
  27. Streisinger G., Okada Y., Emrich J., Newton J., Tsugita A., Terzaghi E., Inouye M. Frameshift mutations and the genetic code. This paper is dedicated to Professor Theodosius Dobzhansky on the occasion of his 66th birthday. Cold Spring Harb Symp Quant Biol. 1966;31:77–84. doi: 10.1101/sqb.1966.031.01.014. [DOI] [PubMed] [Google Scholar]
  28. Taketo A. Sensitivity of Escherichia coli to viral nucleic acid. V. Competence of calcium-treated cells. J Biochem. 1972 Oct;72(4):973–979. doi: 10.1093/oxfordjournals.jbchem.a129988. [DOI] [PubMed] [Google Scholar]
  29. Topal M. D., Fresco J. R. Complementary base pairing and the origin of substitution mutations. Nature. 1976 Sep 23;263(5575):285–289. doi: 10.1038/263285a0. [DOI] [PubMed] [Google Scholar]
  30. 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]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES