Genetic Engineering of Rice for Resistance to Sheath Blight

Abstract

A 1.1 kb rice genomic DNA fragment, containing a chitinase gene under the control of the CaMV 35S promoter, was cloned into the rice transformation vector pGL2. After transformation of Indica rice protoplasts in the presence of polyethyleneglycol, plants were regenerated. The presence of the chimeric chitinase gene in T0 and T1 transgenic rice plants was detected by Southern blot analysis. Western blot analysis of transgenic plants and their progeny revealed the presence of two proteins with apparent molecular weights of 30 and 35 kDa that reacted with the chitinase antibody. Progeny from the chitinase-positive plants were tested for their resistance to the sheath blight pathogen, Rhizoctonia solani. The degree of resistance displayed by the transgenic plants to this pathogen correlated with the level of chitinase expression.

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References

  1. 1

    Collinge, D.B. and Slusarenko, A.J. 1987. Plant gene expression in response to pathogens. Plant Mol. Biol. 9: 389–410.

    CAS  Article  Google Scholar 

  2. 2

    Linthorst, H. J. M. 1991. Pathogenesis-related proteins of plants. Critical Reviews in Plant Sciences 10: 123–150.

    CAS  Article  Google Scholar 

  3. 3

    Boiler, T., Gehri, A., Mauch, F. and Vogeli, U. 1983. Chitinase in bean leaves; induction by ethylene, purification, properties, and possible function. Planta 157: 22–31.

    Article  Google Scholar 

  4. 4

    Vogeli-Lange, R., Hansen-Gehri, A., Boiler, T. and Meins, F. 1988. Induction of the defense-related glucanohydrolases, β-1,3-glucanase and chitinase, by tobacco mosaic virus infection of tobacco leaves. Plant Sci. 54: 171–176.

    Article  Google Scholar 

  5. 5

    Neale, A.D., Wahleithner, J.A., Lund, M., Bonnett, H.T., Kelly, A., Meeks-Wagner, D.R., Peacock, W.J. and Dennis, E.S. 1990. Chitinase, β-1, 3-glucanase, osmotin, and extensin are expressed in tobacco explants during flower formation. The Plant Cell 2: 673–84.

    CAS  PubMed  PubMed Central  Google Scholar 

  6. 6

    Swegle, M., Kramer, K.J. and Muthukrishnan, S. 1992. Properties of barley seed chitinases and release of embryo-associated isoforms during early stages of imbibition. Plant Physiol. 99: 1009–1014.

    CAS  Article  Google Scholar 

  7. 7

    Boller, T. 1985. Induction of hydrolases as a defense reaction against pathogens, p. 247–262. In: Cellular and Molecular Biology of Plant Stress. Key, J. L. and Kosuge, T. (Eds.). Alan R. Liss, New York.

    Google Scholar 

  8. 8

    Schlumbaum, A., Mauch, F., Vogeli, U. and Boller, T. 1986 Plant chitinases are potent inhibitors of fungal growth. Nature 324: 365–367.

    CAS  Article  Google Scholar 

  9. 9

    Roberts, W.K. and Selitrennikoff, C. P. 1988. Plant and bacterial chitinases differ in antifungal activity. J. of Gen. Microbiol. 134: 169–76.

    CAS  Google Scholar 

  10. 10

    Broglie, K.E., Gaynor,J.J. and Broglie, R.M. 1986. Ethylene-regulated gene expression: Molecular cloning of the genes encoding an endochitinase from Phaseolus vulgaris. Proc. Natl. Acad. Sci. USA 83: 6820–6824.

    CAS  Article  Google Scholar 

  11. 11

    Brederode, F.T., Linthorst, H.J.M. and Bol, J.F. 1991. Differential induction of acquired resistance and PR gene expression in tobacco by virus infection, ethephon treatment, UV light and wounding. Plant Mol. Biol. 17: 1117–1125.

    CAS  Article  Google Scholar 

  12. 12

    Gaffney, T., Friedrich, L., Vernooij, B., Negrotto, D., Nye, G., Uknes, S., Ward, E., Kessmann, H. and Ryals, J. 1993. Requirement of salicylic acid for the induction of systemic acquired resistance. Science 261: 754–756.

    CAS  Article  Google Scholar 

  13. 13

    Broglie, K., Chet, I., Holliday, M., Cressman, R., Biddle, P., Knowlton, S., Mauvais, J. and Broglie, R.M. 1991. Transgenic plants with enhanced resistance to the fungal pathogen Rhizoctonia solani. Science 254: 1194–1197.

    CAS  Article  Google Scholar 

  14. 14

    Vierheilig, H., Alt, M., Neuhas, J.-M., Boller, T. and Wiemken, A. 1993. Colonization of transgenic N. sylvestris plants, expressing different forms of N. tabacum chitinase, by the root pathogen Rhizoctonia solani and by the mycorrhizal symbiont Glomus mosseau. Molec. Plant Microbe Interactions 6: 261–264.

    CAS  Article  Google Scholar 

  15. 15

    Toyoda, H., Matsuda, Y., Yamaga, T., Ikeda, S., Morita, M., Tamai, T. and Ouchi, S. 1991. Suppression of the powdery mildew pathogen by chitinase microinjected into barley coleoptile epidermal cells. Plant Cell Reports 10: 217–220.

    CAS  PubMed  Google Scholar 

  16. 16

    Ren, Y. and West, C.A. 1992. Elicitation of diterpene biosynthesis in rice (Oryza saliva L.) by chitin. Plant Physiol. 99: 1169–1178.

    CAS  Article  Google Scholar 

  17. 17

    Huang, J., Wen, L., Swegle, M., Tran, H., Thin, T.H., Naylor, H.M., Muthukrishnan, S. and Reeck, G.R. 1991. Nucleotide sequence of a rice genomic clone that encodes a class I endochitinase. Plant Mol. Biol. 16: 479–480.

    CAS  Article  Google Scholar 

  18. 18

    Boiler, T. 1992. Biochemical analysis of chitinases and β-1,3-glucanases, p. 22–30. In: Molecular Plant Pathology, Vol.II.Gurr,S.J.,McPherson,M.J. and Bowles,D.J.(Eds.). Oxford Univ. Press, UK.

    Google Scholar 

  19. 19

    Ou, S.H. 1985. Rice Diseases. Second Ed, p. 280–282. Commonwealth Mycological Institute Publication. Kew, Surrey, UK.

    Google Scholar 

  20. 20

    Fujimoto, H., Itoh, K., Kyojuka, J. and Shimamoto, K. 1993. Insect resistant rice generated by introduction of a modified δ-endotoxin gene of Bacillus thuringiensis. Bio/Technology 11: 1151–1155.

    CAS  PubMed  Google Scholar 

  21. 21

    Datta, S.K., Peterhans, A., Datta, K. and Potrykus, I. 1990. Genetically engineered fertile Indica-rice plants recovered from protoplasts. Bio/Technology 8: 736–740.

    CAS  Google Scholar 

  22. 22

    Datta, S.K., Datta, K., Soltanifar, N., Donn, G. and Potrykus, I. 1992. Herbicide-resistant Indica rice plants from IRRI breeding line IR72 after PEG-mediated transformation of protoplasts. Plant Mol Biol. 20: 619–629.

    CAS  Article  Google Scholar 

  23. 23

    Vasil, I.K. 1990. Transgenic cereals becoming a reality. Bio/Technology 8: 797.

    Google Scholar 

  24. 24

    Peng, J., Kononowicz, H. and Hodges, T.K. 1992. Transgenic Indica rice plants. Theor. Appl. Genet. 83: 855–863.

    CAS  Article  Google Scholar 

  25. 25

    Christou, P., Tameria, L.F. and Kofron, M. 1991. Production of transgenic rice (Oryza sativa) plants from agronomically important Indica and Japonica varieties via electric discharge particle acceleration of exogenous DNA into immature zygotic embryos. Bio/Technology 9: 957–962.

    Article  Google Scholar 

  26. 26

    Vasil, V., Srivastava, V., Castillo, A.M., Fromm, M.E. and Vasil, I.K. 1993. Rapid production of transgenic wheat plants by direct bombardment of cultured immature embryos. Bio/Technology 11: 1553–1558.

    Google Scholar 

  27. 27

    Koziel, M.G., Beland, G.L., Bowman, C., Carozzi, N.B., Crenshaw, R., Crossland, L., Dawson, J., Desai, N., Hill, M., Kadwell, S., Launis, K., Lewis, K., Maddox, D., McPherson, K., Meghji, M.R., Merlin, E., Rhodes, R., Warren, G.W., Wright, M. and Evola, S.V. 1993. Field performance of elite transgenic maize plants expressing an insecticidal protein derived from Bacillus thuringiensis. Bio/Technology 11: 194–200.

    CAS  Google Scholar 

  28. 28

    Wan, Y. and Lemaux, P. 1994. Generation of large numbers of independently transformed fertile barley plants. Plant Physiol. 104: 37–48.

    CAS  Article  Google Scholar 

  29. 29

    Zhu, Q., Maher, E.A., Masoud, S., Dixon, R.A. and Lamb, C. 1994. Enhanced protection against fungal attack by constitutive co-expression of chitinase and glucanase genes in transgenic tobacco. Bio/Technology 12: 807–812.

    CAS  Google Scholar 

  30. 30

    Anuratha, C.S., Huang, J.-K., Pingali, A. and Muthukrishnan, S. 1992. Isolation and characterization of a chitinase and its cDNA clone from rice. J. Plant Biochem. & Biotech. 1: 5–10.

    CAS  Article  Google Scholar 

  31. 31

    Hart, C.M., Fischer, B., Neuhaus, J.-M. and Meins, F. 1992. Regulated inac-tivation of homologous gene expression in transgenic Nicotiana sylvestris plants containing a defense-related tobacco chitinase gene. Mol Gen. Genet. 235: 179–188.

    CAS  Article  Google Scholar 

  32. 32

    Joshi, C.P. 1987. An inspection of the domain between putative TATA-box and translation start in seventy-nine plant genes. Nucl. Acids Res. 15: 6643–6653.

    CAS  Article  Google Scholar 

  33. 33

    Fromm, M.E., Taylor, L.P. and Walbot, V. 1986. Stable transformation of maize after gene transfer by electroporation. Nature 319: 791–793.

    CAS  Article  Google Scholar 

  34. 34

    Bilang, R., Iida, S., Peterhans, A., Potrykus, I. and Paszkowski, J. 1991. The 3′-terminal region of the hygromycin-B-resistance gene is important for its activity in Escherichia coli and Nicotiana tabacum. Gene 100: 247–250.

    CAS  Article  Google Scholar 

  35. 35

    Murashige, T. and Skoog, F. 1962. A revised medium for a rapid growth and bioassay with tobacco tissue culture. Physiol. Plant. 15: 473–479.

    CAS  Article  Google Scholar 

  36. 36

    Ohira, K., Ojima, K. and Fijiwara, A. 1973. Studies on the nutrition of rice cell culture I. A simple, defined medium for rapid growth in suspension culture. Plant Cell Physiol. 14: 1113–1121.

    CAS  Google Scholar 

  37. 37

    Blackshear, P. J. 1984. Systems for polyacrylamide gel electrophoresis. Methods in Enzymol. 104: 237–257.

    CAS  Article  Google Scholar 

  38. 38

    Winston, S., Fuller, S. and Hurrel, J. 1987. Western blotting, p. 10.8.1–10.8.6. In: Current Protocols in Molecular Biology, John Wiley, NY.

    Google Scholar 

  39. 39

    Murray, M.G. and Thompson, W.F. 1980. Rapid isolation of high molecular weight plant DNA. Nucl. Acids Res. 8: 4321– 4325.

    CAS  Article  Google Scholar 

  40. 40

    Sambrook, J., Fritsch, E.F. and Maniatis, T. 1989. Molecular Cloning: A Laboratory Manual. Cold Spring Harbor, NY.

    Google Scholar 

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Correspondence to S. Muthukrishnan.

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Lin, W., Anuratha, C., Datta, K. et al. Genetic Engineering of Rice for Resistance to Sheath Blight. Nat Biotechnol 13, 686–691 (1995). https://doi.org/10.1038/nbt0795-686

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