Skip to main content
Genetics logoLink to Genetics
. 1990 Apr;124(4):833–843. doi: 10.1093/genetics/124.4.833

Pathways of Transformation in Ustilago Maydis Determined by DNA Conformation

S Fotheringham 1, W K Holloman 1
PMCID: PMC1203975  PMID: 2323553

Abstract

Ustilago maydis was transformed by plasmids bearing a cloned, selectable gene but lacking an autonomously replicating sequence. Transformation was primarily through integration at nonhomologous loci when the plasmid DNA was circular. When the DNA was made linear by cleavage within the cloned gene, the spectrum of integration events shifted from random to targeted recombination at the resident chromosomal allele. In a large fraction of the transformants obtained using linear DNA, the plasmid DNA was not integrated but was maintained in an extrachromosomal state composed of a concatameric array of plasmid units joined end-to-end. The results suggest the operation of several pathways for transformation in U. maydis, and that DNA conformation at the time of transformation governs choice of pathways.

Full Text

The Full Text of this article is available as a PDF (5.1 MB).

Selected References

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

  1. Banks G. R., Taylor S. Y. Cloning of the PYR3 gene of Ustilago maydis and its use in DNA transformation. Mol Cell Biol. 1988 Dec;8(12):5417–5424. doi: 10.1128/mcb.8.12.5417. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Beverley S. M. Characterization of the 'unusual' mobility of large circular DNAs in pulsed field-gradient electrophoresis. Nucleic Acids Res. 1988 Feb 11;16(3):925–939. doi: 10.1093/nar/16.3.925. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Case M. E. Genetical and molecular analyses of qa-2 transformants in Neurospora crassa. Genetics. 1986 Jul;113(3):569–587. doi: 10.1093/genetics/113.3.569. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Celniker S. E., Sweder K., Srienc F., Bailey J. E., Campbell J. L. Deletion mutations affecting autonomously replicating sequence ARS1 of Saccharomyces cerevisiae. Mol Cell Biol. 1984 Nov;4(11):2455–2466. doi: 10.1128/mcb.4.11.2455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chang X. B., Wilson J. H. Modification of DNA ends can decrease end joining relative to homologous recombination in mammalian cells. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4959–4963. doi: 10.1073/pnas.84.14.4959. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chu G., Vollrath D., Davis R. W. Separation of large DNA molecules by contour-clamped homogeneous electric fields. Science. 1986 Dec 19;234(4783):1582–1585. doi: 10.1126/science.3538420. [DOI] [PubMed] [Google Scholar]
  7. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  8. Gritz L., Davies J. Plasmid-encoded hygromycin B resistance: the sequence of hygromycin B phosphotransferase gene and its expression in Escherichia coli and Saccharomyces cerevisiae. Gene. 1983 Nov;25(2-3):179–188. doi: 10.1016/0378-1119(83)90223-8. [DOI] [PubMed] [Google Scholar]
  9. Heyer W. D., Sipiczki M., Kohli J. Replicating plasmids in Schizosaccharomyces pombe: improvement of symmetric segregation by a new genetic element. Mol Cell Biol. 1986 Jan;6(1):80–89. doi: 10.1128/mcb.6.1.80. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hightower R. C., Metge D. W., Santi D. V. Plasmid migration using orthogonal-field-alternation gel electrophoresis. Nucleic Acids Res. 1987 Oct 26;15(20):8387–8398. doi: 10.1093/nar/15.20.8387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hinnen A., Hicks J. B., Fink G. R. Transformation of yeast. Proc Natl Acad Sci U S A. 1978 Apr;75(4):1929–1933. doi: 10.1073/pnas.75.4.1929. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hoffman C. S., Winston F. A ten-minute DNA preparation from yeast efficiently releases autonomous plasmids for transformation of Escherichia coli. Gene. 1987;57(2-3):267–272. doi: 10.1016/0378-1119(87)90131-4. [DOI] [PubMed] [Google Scholar]
  13. Holliday R. Altered recombination frequencies in radiation sensitivie strains of Ustilago. Mutat Res. 1967 May-Jun;4(3):275–288. doi: 10.1016/0027-5107(67)90022-x. [DOI] [PubMed] [Google Scholar]
  14. Holliday R., Halliwell R. E., Evans M. W., Rowell V. Genetic characterization of rec-1, a mutant of Ustilago maydis defective in repair and recombination. Genet Res. 1976 Jun;27(3):413–453. doi: 10.1017/s0016672300016621. [DOI] [PubMed] [Google Scholar]
  15. Holliday R. Studies on mitotic gene conversion in Ustilago. Genet Res. 1966 Dec;8(3):323–337. doi: 10.1017/s0016672300010181. [DOI] [PubMed] [Google Scholar]
  16. Kato S., Anderson R. A., Camerini-Otero R. D. Foreign DNA introduced by calcium phosphate is integrated into repetitive DNA elements of the mouse L cell genome. Mol Cell Biol. 1986 May;6(5):1787–1795. doi: 10.1128/mcb.6.5.1787. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kearsey S. Structural requirements for the function of a yeast chromosomal replicator. Cell. 1984 May;37(1):299–307. doi: 10.1016/0092-8674(84)90326-x. [DOI] [PubMed] [Google Scholar]
  18. Lilley D. M. In vivo consequences of plasmid topology. Nature. 1981 Jul 23;292(5821):380–382. doi: 10.1038/292380a0. [DOI] [PubMed] [Google Scholar]
  19. Lin F. L., Sperle K., Sternberg N. Model for homologous recombination during transfer of DNA into mouse L cells: role for DNA ends in the recombination process. Mol Cell Biol. 1984 Jun;4(6):1020–1034. doi: 10.1128/mcb.4.6.1020. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Maundrell K., Hutchison A., Shall S. Sequence analysis of ARS elements in fission yeast. EMBO J. 1988 Jul;7(7):2203–2209. doi: 10.1002/j.1460-2075.1988.tb03059.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. McClintock B. The Behavior in Successive Nuclear Divisions of a Chromosome Broken at Meiosis. Proc Natl Acad Sci U S A. 1939 Aug;25(8):405–416. doi: 10.1073/pnas.25.8.405. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Orr-Weaver T. L., Szostak J. W., Rothstein R. J. Yeast transformation: a model system for the study of recombination. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6354–6358. doi: 10.1073/pnas.78.10.6354. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Orr-Weaver T. L., Szostak J. W. Yeast recombination: the association between double-strand gap repair and crossing-over. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4417–4421. doi: 10.1073/pnas.80.14.4417. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Paietta J. V., Marzluf G. A. Gene disruption by transformation in Neurospora crassa. Mol Cell Biol. 1985 Jul;5(7):1554–1559. doi: 10.1128/mcb.5.7.1554. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Palzkill T. G., Newlon C. S. A yeast replication origin consists of multiple copies of a small conserved sequence. Cell. 1988 May 6;53(3):441–450. doi: 10.1016/0092-8674(88)90164-x. [DOI] [PubMed] [Google Scholar]
  26. ROMAN H., JACOB F. A comparison of spontaneous and ultraviolet-induced allelic recombination with reference to the recombination of outside markers. Cold Spring Harb Symp Quant Biol. 1958;23:155–160. doi: 10.1101/sqb.1958.023.01.019. [DOI] [PubMed] [Google Scholar]
  27. Ray A., Siddiqi I., Kolodkin A. L., Stahl F. W. Intra-chromosomal gene conversion induced by a DNA double-strand break in Saccharomyces cerevisiae. J Mol Biol. 1988 May 20;201(2):247–260. doi: 10.1016/0022-2836(88)90136-2. [DOI] [PubMed] [Google Scholar]
  28. Reed K. C., Mann D. A. Rapid transfer of DNA from agarose gels to nylon membranes. Nucleic Acids Res. 1985 Oct 25;13(20):7207–7221. doi: 10.1093/nar/13.20.7207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Robins D. M., Ripley S., Henderson A. S., Axel R. Transforming DNA integrates into the host chromosome. Cell. 1981 Jan;23(1):29–39. doi: 10.1016/0092-8674(81)90267-1. [DOI] [PubMed] [Google Scholar]
  30. Roman H., Fabre F. Gene conversion and associated reciprocal recombination are separable events in vegetative cells of Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1983 Nov;80(22):6912–6916. doi: 10.1073/pnas.80.22.6912. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Song K. Y., Chekuri L., Rauth S., Ehrlich S., Kucherlapati R. Effect of double-strand breaks on homologous recombination in mammalian cells and extracts. Mol Cell Biol. 1985 Dec;5(12):3331–3336. doi: 10.1128/mcb.5.12.3331. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Szostak J. W., Orr-Weaver T. L., Rothstein R. J., Stahl F. W. The double-strand-break repair model for recombination. Cell. 1983 May;33(1):25–35. doi: 10.1016/0092-8674(83)90331-8. [DOI] [PubMed] [Google Scholar]
  33. Thomas K. R., Folger K. R., Capecchi M. R. High frequency targeting of genes to specific sites in the mammalian genome. Cell. 1986 Feb 14;44(3):419–428. doi: 10.1016/0092-8674(86)90463-0. [DOI] [PubMed] [Google Scholar]
  34. Tsukuda T., Carleton S., Fotheringham S., Holloman W. K. Isolation and characterization of an autonomously replicating sequence from Ustilago maydis. Mol Cell Biol. 1988 Sep;8(9):3703–3709. doi: 10.1128/mcb.8.9.3703. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Umek R. M., Kowalski D. The ease of DNA unwinding as a determinant of initiation at yeast replication origins. Cell. 1988 Feb 26;52(4):559–567. doi: 10.1016/0092-8674(88)90469-2. [DOI] [PubMed] [Google Scholar]
  36. Vollrath D., Davis R. W., Connelly C., Hieter P. Physical mapping of large DNA by chromosome fragmentation. Proc Natl Acad Sci U S A. 1988 Aug;85(16):6027–6031. doi: 10.1073/pnas.85.16.6027. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Genetics are provided here courtesy of Oxford University Press

RESOURCES