1887

Abstract

Four ribozyme and antisense genes targeting citrus exocortis viroid (CEVd) positive- and negative-strand RNA molecules were constructed and used to transform the tomato cv. UC82B. The tomato is a readily transformable plant and will support replication of CEVd following mechanical inoculation. The ribozyme genes contained three hammerhead catalytic motifs with long hybridizing arms and synthetic RNA transcripts were shown to cleave the target CEVd RNA molecule . Homozygous transgenic plants were produced from independent transformants expressing either ribozymes or antisense constructs. Inoculation of transgenic seedlings expressing antisense constructs targeting the negative-strand CEVd RNA molecule with CEVd resulted in a moderate reduction in the accumulation of CEVd RNA. In contrast, similarly inoculated transgenic plants expressing constructs targeting the positive-strand CEVd RNA molecule resulted in an increase in the rate of CEVd RNA accumulation. Addition of the ribozyme motifs to the antisense genes did not enhance their efficiency in the suppression of viroid replication and a moderation or elimination of the observed antisense effects was seen in plants expressing the corresponding catalytic RNA-encoding genes.

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1995-07-01
2024-03-28
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References

  1. An G., Watson B. D., Stachel S., Gordon M. P., Nester E. W. 1985; New cloning vehicles for transformation of higher plants. EMBO Journal 4:277–284
    [Google Scholar]
  2. Beachy R. N., Loesch-Fries S., Tumer N. E. 1990; Coat protein-mediated resistance against virus infection. Annual Review of Phytopathology 28:451–474
    [Google Scholar]
  3. Cuozzo M., O’Connell K. M., Kaniewski W., Fang R.-X., Chua N.-H., Tumer N. E. 1988; Viral protection in transgenic tobacco plants expressing the cucumber mosaic virus coat protein or its antisense RNA. Bio/Technology 6:549–557
    [Google Scholar]
  4. Diener T. O. 1987 The Plant Viruses vol 2 Edited by Regenmortel M. H. V., Fraenkel-Conrat H. Boca Raton: CRC Press;
    [Google Scholar]
  5. Edington B. V., Nelson R. S. 1992; Utilisation of ribozymes in plants: plant viral resistance. In Gene Regulation: Biology of Antisense RNA and DNA pp 209–222 Edited by Erickson R. P., Izant J. G. New York: Raven Press;
    [Google Scholar]
  6. Fillatti J. J., Kiser J., Rose R., Comai L. 1987; Efficient transfer of a glyphosphate tolerance gene into tomato using a binary Agrobacterium tumefaciens vector. Bio/ Technology 5:726–730
    [Google Scholar]
  7. Garfinkel D. J., Nester A. W. 1980; Agrobacterium tumefaciens mutants affected in crown gall tumorigenesis and octopine catabolism. Journal of Bacteriology 144:732–743
    [Google Scholar]
  8. Gerlach W. L., Bedbrook J. R. 1979; Cloning and characterisation of ribosomal RNA genes from wheat and barley. Nucleic Acids Research 7:1869–1885
    [Google Scholar]
  9. Golemboski D. B., Lomonossoff G. P., Zaitlin M. 1990; Plants transformed with a tobacco mosaic virus non-structural gene sequence are resistant to the virus. Proceedings of the National Academy of Sciences, USA 87:6311–6315
    [Google Scholar]
  10. Haseloff J., Gerlach W. L. 1988; Simple RNA enzymes with new and highly specific endonuclease activities. Nature 334:585–591
    [Google Scholar]
  11. Hemenway C., Fang R.-X., Kaniewski W., Chua N.-H., Tumer N.-E. 1988; Analysis of the mechanism of protection in transgenic plants expressing the potato virus X coat protein or its antisense. EMBO Journal 1:1273–1280
    [Google Scholar]
  12. Hillman B. I., Carrington J. C., Morris T. J. 1987; A defective interfering RNA that contains a mosaic of a plant virus genome. Cell 51:427–433
    [Google Scholar]
  13. Hutchins C. J., Keese P., Visvader J. E., Rathjen P. D., McInnes J. L., Symons R. H. 1985; Comparison of multimeric plus and minus forms of viroids and virusoids. Plant Molecular Biology 4:293–304
    [Google Scholar]
  14. Kawchuk L. M., Martin R. R., McPherson J. 1991; Sense and antisense RNA-mediated resistance to potato leafroll virus in Russet Burbank potato plants. Molecular Plant-Microbe Interactions 4:247–253
    [Google Scholar]
  15. Kunkel T. A., Roberts J. D., Zakour R. A. 1987; Rapid and efficient specific mutagenesis without phenotypic selection. Methods in Enzymology 154:267–382
    [Google Scholar]
  16. MacFarlane S. A., Davies J. W. 1992; Plants transformed with a region of the 201-kilodalton replicase gene from pea early browning virus RNA1 are resistant to virus infection. Proceedings of the National Academy of Sciences, USA 89:5829–5833
    [Google Scholar]
  17. McDonnell R. E., Clark R. D., Smith W. A., Hinchee M. A. 1987; A simplified method for the detection of neomycin phosphotransferase II activity in transformed plant tissues. Plant Molecular Biology Reporter 5:380–386
    [Google Scholar]
  18. Melton D. A., Krieg P. A., Rebagliati M. R., Maniatis T., Zinn K., Green M. R. 1984; Efficient in vitro synthesis of biologically active RNA and RNA hybridisation probes from plasmids containing a bacteriophage SP6 promoter. Nucleic Acids Research 12:7035–7056
    [Google Scholar]
  19. Palukaitis P. 1987; Potato spindle tuber viroid: investigation of the long-distance, intra-plant transport route. Virology 158:239–241
    [Google Scholar]
  20. Riesner D. 1987; Viroid function: subcellular location and in situ association with cellular components. In The Viroids pp 99–116 Edited by Diener T. O. New York: Plenum Press;
    [Google Scholar]
  21. Rigden J. E., Rezaian M. A. 1992; In vitro synthesis of an infectious viroid: analysis of the infectivity of monomeric linear CEV. Virology 186:201–206
    [Google Scholar]
  22. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning: A Laboratory Manual New York: Cold Spring Harbor Laboratory;
    [Google Scholar]
  23. Sanford J. C., Johnston S. A. 1985; The concept of pathogen derived resistance: deriving resistance genes from the parasites own genome. Journal of Theoretical Biology 113:395–405
    [Google Scholar]
  24. Sanger F., Nicklen S., Coulson A. R. 1977; DNA sequencing with chain-terminating inhibitors. Proceedings of the National Academy of Sciences, USA 74:5463–5467
    [Google Scholar]
  25. Sänger H. L. 1987; Viroid replication. In The Viroids pp 117–166 Edited by Diener T. O. New York: Plenum Press;
    [Google Scholar]
  26. Sänger H. L., Klotz G., Riesner D., Gross H. J., Kleinschmidt A. K. 1976; Viroids are single-stranded covalently closed circular RNA molecules existing as highly base-paired rod-like structures. Proceedings of the National Academy of Sciences, USA 73:3852–3856
    [Google Scholar]
  27. Steinecke P., Herget T., Shreir P. H. 1992; Expression of a chimaeric ribozyme gene results in endonucleolytic cleavage of target mRNA and concomitant reduction of gene expression in vivo . EMBO Journal 11:1525–1530
    [Google Scholar]
  28. Symons R. H. 1990; The fascination of low molecular weight pathogenic RNAs. Seminars in Virology 1:75–81
    [Google Scholar]
  29. Thomas B., Pratt D. 1981; Efficient hybridisation between Lycopersicon esculentum and L. peruvianum via embryo callus. Theoretical and Applied Genetics 59:215–219
    [Google Scholar]
  30. Van der Krol A. R., Mol J. N. M., Stuitje A. R. 1988a; Antisense genes in plants: an overview. Gene 72:45–50
    [Google Scholar]
  31. Van der Krol A. R., Lenting P. E., Veenstra J., Van Der Meer I. M., Koes R. E., Gerats A. G. M., Mol J. N. M., Stuitje A. R. 1988b; An anti-sense chalcone synthase gene in transgenic plants inhibits flower pigmentation. Nature 333:866–869
    [Google Scholar]
  32. Visvader J. E., Gould A. R., Breuning G. E., Symons R. H. 1982; Citrus exocortis viroid: nucleotide sequence and secondary structure of an Australian isolate. FEBS Letters 137:288–292
    [Google Scholar]
  33. Walker J. C., Howard E. A., Dennis E. S., Peacock W. J. 1987; DNA sequences required for anaerobic expression of the maize adh l gene. Proceedings of the National Academy of Sciences, USA 84:6624–6628
    [Google Scholar]
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