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. 1987 Mar;169(3):1046–1055. doi: 10.1128/jb.169.3.1046-1055.1987

Processing of the T-DNA of Agrobacterium tumefaciens generates border nicks and linear, single-stranded T-DNA.

L M Albright, M F Yanofsky, B Leroux, D Q Ma, E W Nester
PMCID: PMC211899  PMID: 3029014

Abstract

Transfer and integration of a defined region (T-DNA) of the tumor-inducing (Ti) plasmid of Agrobacterium tumefaciens is essential for tumor formation. We used a physical assay to study structural changes induced in Agrobacterium T-DNA by cocultivation with plant cells. We show that nicks are introduced at unique, identical locations in each of the 24-base-pair imperfect direct repeats which flank the T-DNA and present evidence that a linear, single-stranded molecule is generated. We propose that these changes result from processing of the T-DNA for transfer and that they occur by a mechanism similar to DNA processing during conjugative DNA transfer between bacteria.

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Selected References

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

  1. Alt-Moerbe J., Rak B., Schröder J. A 3.6-kbp segment from the vir region of Ti plasmids contains genes responsible for border-sequence-directed production of T region circles in E. coli. EMBO J. 1986 Jun;5(6):1129–1135. doi: 10.1002/j.1460-2075.1986.tb04337.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. An G. High efficiency transformation of cultured tobacco cells. Plant Physiol. 1985 Oct;79(2):568–570. doi: 10.1104/pp.79.2.568. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Beringer J. E. R factor transfer in Rhizobium leguminosarum. J Gen Microbiol. 1974 Sep;84(1):188–198. doi: 10.1099/00221287-84-1-188. [DOI] [PubMed] [Google Scholar]
  4. Bevan M. W., Chilton M. D. T-DNA of the Agrobacterium Ti and Ri plasmids. Annu Rev Genet. 1982;16:357–384. doi: 10.1146/annurev.ge.16.120182.002041. [DOI] [PubMed] [Google Scholar]
  5. 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]
  6. 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]
  7. Caplan A., Herrera-Estrella L., Inzé D., Van Haute E., Van Montagu M., Schell J., Zambryski P. Introduction of genetic material into plant cells. Science. 1983 Nov 18;222(4625):815–821. doi: 10.1126/science.222.4625.815. [DOI] [PubMed] [Google Scholar]
  8. Chilton M. D., Currier T. C., Farrand S. K., Bendich A. J., Gordon M. P., Nester E. W. Agrobacterium tumefaciens DNA and PS8 bacteriophage DNA not detected in crown gall tumors. Proc Natl Acad Sci U S A. 1974 Sep;71(9):3672–3676. doi: 10.1073/pnas.71.9.3672. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fromm M. E., Taylor L. P., Walbot V. Stable transformation of maize after gene transfer by electroporation. 1986 Feb 27-Mar 5Nature. 319(6056):791–793. doi: 10.1038/319791a0. [DOI] [PubMed] [Google Scholar]
  10. Gardner R. C., Knauf V. C. Transfer of Agrobacterium DNA to Plants Requires a T-DNA Border But Not the virE Locus. Science. 1986 Feb 14;231(4739):725–727. doi: 10.1126/science.231.4739.725. [DOI] [PubMed] [Google Scholar]
  11. Garfinkel D. J., Simpson R. B., Ream L. W., White F. F., Gordon M. P., Nester E. W. Genetic analysis of crown gall: fine structure map of the T-DNA by site-directed mutagenesis. Cell. 1981 Nov;27(1 Pt 2):143–153. doi: 10.1016/0092-8674(81)90368-8. [DOI] [PubMed] [Google Scholar]
  12. Horsch R. B., Klee H. J. Rapid assay of foreign gene expression in leaf discs transformed by Agrobacterium tumefaciens: Role of T-DNA borders in the transfer process. Proc Natl Acad Sci U S A. 1986 Jun;83(12):4428–4432. doi: 10.1073/pnas.83.12.4428. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Horsch R. B., Klee H. J., Stachel S., Winans S. C., Nester E. W., Rogers S. G., Fraley R. T. Analysis of Agrobacterium tumefaciens virulence mutants in leaf discs. Proc Natl Acad Sci U S A. 1986 Apr;83(8):2571–2575. doi: 10.1073/pnas.83.8.2571. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Jen G. C., Chilton M. D. Activity of T-DNA borders in plant cell transformation by mini-T plasmids. J Bacteriol. 1986 May;166(2):491–499. doi: 10.1128/jb.166.2.491-499.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Jen G. C., Chilton M. D. The right border region of pTiT37 T-DNA is intrinsically more active than the left border region in promoting T-DNA transformation. Proc Natl Acad Sci U S A. 1986 Jun;83(11):3895–3899. doi: 10.1073/pnas.83.11.3895. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Joos H., Timmerman B., Montagu M. V., Schell J. Genetic analysis of transfer and stabilization of Agrobacterium DNA in plant cells. EMBO J. 1983;2(12):2151–2160. doi: 10.1002/j.1460-2075.1983.tb01716.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Klee H. J., White F. F., Iyer V. N., Gordon M. P., Nester E. W. Mutational analysis of the virulence region of an Agrobacterium tumefaciens Ti plasmid. J Bacteriol. 1983 Feb;153(2):878–883. doi: 10.1128/jb.153.2.878-883.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Knauf V. C., Nester E. W. Wide host range cloning vectors: a cosmid clone bank of an Agrobacterium Ti plasmid. Plasmid. 1982 Jul;8(1):45–54. doi: 10.1016/0147-619x(82)90040-3. [DOI] [PubMed] [Google Scholar]
  19. Melton D. A., Krieg P. A., Rebagliati M. R., Maniatis T., Zinn K., Green M. R. Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter. Nucleic Acids Res. 1984 Sep 25;12(18):7035–7056. doi: 10.1093/nar/12.18.7035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Peacock A. C., Dingman C. W. Molecular weight estimation and separation of ribonucleic acid by electrophoresis in agarose-acrylamide composite gels. Biochemistry. 1968 Feb;7(2):668–674. doi: 10.1021/bi00842a023. [DOI] [PubMed] [Google Scholar]
  21. Peralta E. G., Ream L. W. T-DNA border sequences required for crown gall tumorigenesis. Proc Natl Acad Sci U S A. 1985 Aug;82(15):5112–5116. doi: 10.1073/pnas.82.15.5112. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Richardson J. P. Attachment of nascent RNA molecules to superhelical DNA. J Mol Biol. 1975 Nov 5;98(3):565–579. doi: 10.1016/s0022-2836(75)80087-8. [DOI] [PubMed] [Google Scholar]
  23. 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]
  24. Simpson R. B., O'Hara P. J., Kwok W., Montoya A. L., Lichtenstein C., Gordon M. P., Nester E. W. DNA from the A6S/2 crown gall tumor contains scrambled Ti-plasmid sequences near its junctions with plant DNA. Cell. 1982 Jul;29(3):1005–1014. doi: 10.1016/0092-8674(82)90464-0. [DOI] [PubMed] [Google Scholar]
  25. Stachel S. E., Nester E. W. The genetic and transcriptional organization of the vir region of the A6 Ti plasmid of Agrobacterium tumefaciens. EMBO J. 1986 Jul;5(7):1445–1454. doi: 10.1002/j.1460-2075.1986.tb04381.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Stachel S. E., Nester E. W., Zambryski P. C. A plant cell factor induces Agrobacterium tumefaciens vir gene expression. Proc Natl Acad Sci U S A. 1986 Jan;83(2):379–383. doi: 10.1073/pnas.83.2.379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Thomas M., White R. L., Davis R. W. Hybridization of RNA to double-stranded DNA: formation of R-loops. Proc Natl Acad Sci U S A. 1976 Jul;73(7):2294–2298. doi: 10.1073/pnas.73.7.2294. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Van Montagu M., Schell J. The Ti plasmids of Agrobacterium. Curr Top Microbiol Immunol. 1982;96:237–254. doi: 10.1007/978-3-642-68315-2_13. [DOI] [PubMed] [Google Scholar]
  29. Wang K., Herrera-Estrella L., Van Montagu M., Zambryski P. Right 25 bp terminus sequence of the nopaline T-DNA is essential for and determines direction of DNA transfer from agrobacterium to the plant genome. Cell. 1984 Sep;38(2):455–462. doi: 10.1016/0092-8674(84)90500-2. [DOI] [PubMed] [Google Scholar]
  30. Willetts N., Wilkins B. Processing of plasmid DNA during bacterial conjugation. Microbiol Rev. 1984 Mar;48(1):24–41. doi: 10.1128/mr.48.1.24-41.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Yadav N. S., Vanderleyden J., Bennett D. R., Barnes W. M., Chilton M. D. Short direct repeats flank the T-DNA on a nopaline Ti plasmid. Proc Natl Acad Sci U S A. 1982 Oct;79(20):6322–6326. doi: 10.1073/pnas.79.20.6322. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Yanofsky M. F., Porter S. G., Young C., Albright L. M., Gordon M. P., Nester E. W. The virD operon of Agrobacterium tumefaciens encodes a site-specific endonuclease. Cell. 1986 Nov 7;47(3):471–477. doi: 10.1016/0092-8674(86)90604-5. [DOI] [PubMed] [Google Scholar]
  33. Zambryski P., Depicker A., Kruger K., Goodman H. M. Tumor induction by Agrobacterium tumefaciens: analysis of the boundaries of T-DNA. J Mol Appl Genet. 1982;1(4):361–370. [PubMed] [Google Scholar]
  34. Zambryski P., Joos H., Genetello C., Leemans J., Montagu M. V., Schell J. Ti plasmid vector for the introduction of DNA into plant cells without alteration of their normal regeneration capacity. EMBO J. 1983;2(12):2143–2150. doi: 10.1002/j.1460-2075.1983.tb01715.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Zinn K., DiMaio D., Maniatis T. Identification of two distinct regulatory regions adjacent to the human beta-interferon gene. Cell. 1983 Oct;34(3):865–879. doi: 10.1016/0092-8674(83)90544-5. [DOI] [PubMed] [Google Scholar]

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