Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1987 Dec 10;15(23):9807–9823. doi: 10.1093/nar/15.23.9807

Molecular cloning and characterization of the gene encoding the DNA methyltransferase, M.CviBIII, from Chlorella virus NC-1A.

K E Narva 1, D L Wendell 1, M P Skrdla 1, J L Van Etten 1
PMCID: PMC306533  PMID: 3320956

Abstract

The gene encoding the DNA methyltransferase, M.CviBIII, from Chlorella virus NC-1A was cloned and expressed in E. coli plasmid pUC8. Plasmid (pNC-1A.14.8) encoded M.CviBIII methylates adenine in TCGA sequences both in vivo in E. coli and in vitro. Transposon Tn5 mutagenesis localized the M.CviBIII functional domain to a 1.5 kbp region of pNC-1A.14.8 and also indicated that a virus promoter directs transcription of the gene in E. coli. The 2.1 kbp insert containing the M.CviBIII gene was sequenced and a single open reading frame of 1131 bp was identified within the domain determined by Tn5 mutagenesis. When the M.CviBIII gene was fused in-frame with the 19 amino-terminal codons of lacZ a 45 kD polypeptide was identified in maxicells as predicted by the DNA sequence. The M.CviBIII gene was not essential for virus replication since a virus M.CviBIII deletion mutant also replicated in Chlorella.

Full text

PDF
9807

Images in this article

9810Image
on p.9810
9813Image
on p.9813
9817Image
on p.9817
9818Image
on p.9818
9819Image
on p.9819

Selected References

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

  1. Biggin M. D., Gibson T. J., Hong G. F. Buffer gradient gels and 35S label as an aid to rapid DNA sequence determination. Proc Natl Acad Sci U S A. 1983 Jul;80(13):3963–3965. doi: 10.1073/pnas.80.13.3963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Mann M. B., Rao R. N., Smith H. O. Cloning of restriction and modification genes in E. coli: the HbaII system from Haemophilus haemolyticus. Gene. 1978 Apr;3(2):97–112. doi: 10.1016/0378-1119(78)90054-9. [DOI] [PubMed] [Google Scholar]
  3. Marinus M. G., Carraway M., Frey A. Z., Brown L., Arraj J. A. Insertion mutations in the dam gene of Escherichia coli K-12. Mol Gen Genet. 1983;192(1-2):288–289. doi: 10.1007/BF00327681. [DOI] [PubMed] [Google Scholar]
  4. Messing J. New M13 vectors for cloning. Methods Enzymol. 1983;101:20–78. doi: 10.1016/0076-6879(83)01005-8. [DOI] [PubMed] [Google Scholar]
  5. Rosenberg M., Court D. Regulatory sequences involved in the promotion and termination of RNA transcription. Annu Rev Genet. 1979;13:319–353. doi: 10.1146/annurev.ge.13.120179.001535. [DOI] [PubMed] [Google Scholar]
  6. 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]
  7. 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]
  8. Schuster A. M., Burbank D. E., Meister B., Skrdla M. P., Meints R. H., Hattman S., Swinton D., Van Etten J. L. Characterization of viruses infecting a eukaryotic Chlorella-like green alga. Virology. 1986 Apr 15;150(1):170–177. doi: 10.1016/0042-6822(86)90276-x. [DOI] [PubMed] [Google Scholar]
  9. Van Etten J. L., Burbank D. E., Schuster A. M., Meints R. H. Lytic viruses infecting a Chlorella-like alga. Virology. 1985 Jan 15;140(1):135–143. doi: 10.1016/0042-6822(85)90452-0. [DOI] [PubMed] [Google Scholar]
  10. Van Etten J. L., Schuster A. M., Girton L., Burbank D. E., Swinton D., Hattman S. DNA methylation of viruses infecting a eukaryotic Chlorella-like green alga. Nucleic Acids Res. 1985 May 24;13(10):3471–3478. doi: 10.1093/nar/13.10.3471. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  12. Xia Y. N., Burbank D. E., Uher L., Rabussay D., Van Etten J. L. Restriction endonuclease activity induced by PBCV-1 virus infection of a Chlorella-like green alga. Mol Cell Biol. 1986 May;6(5):1430–1439. doi: 10.1128/mcb.6.5.1430. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Xia Y. N., Van Etten J. L. DNA methyltransferase induced by PBCV-1 virus infection of a Chlorella-like green alga. Mol Cell Biol. 1986 May;6(5):1440–1445. doi: 10.1128/mcb.6.5.1440. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Xia Y., Burbank D. E., Van Etten J. L. Restriction endonuclease activity induced by NC-1A virus infection of a Chlorella-like green alga. Nucleic Acids Res. 1986 Aug 11;14(15):6017–6030. doi: 10.1093/nar/14.15.6017. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]
  16. de Bruijn F. J., Lupski J. R. The use of transposon Tn5 mutagenesis in the rapid generation of correlated physical and genetic maps of DNA segments cloned into multicopy plasmids--a review. Gene. 1984 Feb;27(2):131–149. doi: 10.1016/0378-1119(84)90135-5. [DOI] [PubMed] [Google Scholar]

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

RESOURCES