Summary
Mature seed-derived callus from an elite Chinese japonica rice cv. Ewan 5 was cotransformed with two plasmids, pWRG1515 and pRSSGNAl, containing the selectable marker hygromycin phosphotransferase gene (hpt), the reporter β-glucuronidase gene (gusA) and the snowdrop (Galanthus nivalis) lectin gene (gna) via particle bombardment. Thirty-five independent transgenic rice plants were regenerated from 177 bombarded calluses. Eighty-three percent of the transgenic plants contained all three genes, as revealed by Southern blot analysis. Western blot analysis revealed that 23 out of 29 gna-containing transgenic plants expressed Galanthus nivalis agglutinin (GNA) (79%) at various levels, with the highest expression being approximately 0.5% of total soluble protein. Genetic analysis confirmed Mendelian segregation of all three transgenes (gna, hpt and gusA) in the R2 progeny. Amongst the R2 generation two independent homozygous lines were identified that expressed all three transgenes. Insect bioassay and feeding tests showed that these homozygous lines had significant inhibition to rice brown planthopper (Nilaparvata lugens, BPH) by decreasing the survival, overall fecundity of BPH, retarding development, and decreasing the feeding of BPH. These BPH-resistant lines have been incorporated into a rice insect resistance breeding program. This is the first report that homozygous transgenic rice lines expressing GNA, developed by genetic transformation and through genetic analysis-based selection, conferred enhanced resistance to BPH.
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Burkhardt, P. K.; Beyer, P.; Wunn, J.; Kloti, A.; Armstrong, G. A.; Schledz, M.; von Lintig, J.; Potrykus, I.; Transgenic rice (Oryza sativa) endosperm expressing daffodil (Narcissus pseudonarcissus) phytoene synthase accumulates phytoene, a key intermediate of provitamin A biosynthesis. Plant J. 11:1071–1078; 1997.
Chen, L.; Zhang, S.; Beachy, R. N. Fauquet, C. M. A protocol for consistent, large-scale production of fertile transgenic rice plants. Plant Cell Rep. 18:25–31; 1998.
Cheng, X.; Sardana, R.; Kaplan, H.; Altosaar, I. Agrobacterium-transformed rice plants expressing synthetic cryIA(b) and cryIA(c) genes are highly toxic to striped stem borer and yellow stem borer. Proc. Natl Acad. Sci. USA 95:2767–2772; 1998.
Christou, P. Strategies for variety-independent genetic transformation of important cereals, legumes and woody species utilizing particle bombardment. Euphytica 85:13–27; 1995.
Christou, P.; Ford, T. L.; Kofron, M. Production of transgenic rice (Oryza sativa L.) 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; 1991.
Christou, P.; Ford, T. L.; Kofron, M. The development of a varietyindependent gene-transfer method for rice. Trends Biotechnol. 10:239–246; 1992.
Cooley, J.; Ford, T.; Christou, P. Molecular and genetic characterization of elite transgenic rice plants produced by electric discharge particle acceleration. Theor. Appl. Genet. 90:97–104; 1995.
Edwards, K.; Johnstone, C.; Thompson, C. A simple and rapid method for the preparation of plant genomic DNA for PCR analyses. Nucl. Acids Res. 98:1349; 1991.
Goto, F.; Toki, S.; Uchimiya, H. Inheritance of a co-transferred foreign gene in the progenies of transgenic rice plants. Transgenic Res. 2:302–305; 1993.
Hadi, M. Z.; McMullen M. D.; Finer, J. J. Transformation of 12 different plasmids into soybean via particle bombardment. Plant Cell Rep. 15:500–505; 1996.
Hiei, Y.; Ohta, S.; Komari, T.; Kumashiro, T. Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA. Plant J. 6:271–282; 1994.
Ishida, Y.; Saito, H.; Ohta, S.; Hiei, Y.; Komari T.; Kumashiro, T. High efficiency transformation of maize (Zea mays L.) mediated by Agrobacterium tumefaciens. Nature Biotechnol. 14:745–750; 1996.
Jefferson, R. A.; Kavanagh, T. A.; Bevan, M. W. GUS fusions: β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J. 6:3901–3907; 1987.
Kohli, A.; Leech, M.; Vain, P.; Laurie, D. A.; Christou, P. Transgene organization in rice engineered through direct DNA transfer supports a two-phase integration mechanism mediated by the establishment of integration hot-spots. Proc. Natl Acad. Sci. USA 95:7203–7207; 1998.
Maqbool, S. B.; Christou, P. Multiple traits of agronomic importance in transgenic indica rice plants: analysis of transgene integration patterns, expression levels and stability. Mol. Breed. 5:471–480; 1999.
Mochida, O.; Wahyu, A.; Surjani, T. K. Some considerations on screening resistant cultivars/lines of rice plant to the brown planthopper, Nilaparvata lugens (Stal) (Hom., Delphacidae). Los Banos, Philippines: IRRI; 1979: 1–9.
Murashige, T.; Skoog, F. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant. 15:473–497; 1962.
Ou, S. M. Rice diseases, 2nd edn. UK: Commonwealth Agriculture Bureau, Commonwealth Mycological Institute; 1985: 360 pp.
Powell, K. S.; Gatehouse, A. M. R.; Hilder, V. A.; Gatehouse, J. A. Antimetabolic effects of plant lectins and plant and fungal enzymes on the nymphal stages of two important rice pest. Nilaparvata lugens and Nephotettix cinciteps. Entomol. Exp. Appl. 66:119–126; 1993.
Powell, K. S.; Gatehouse, A. M. R.; Hilder, V. A.; Gatehouse, J. A. Antifeedant effects of plant lectins and enzymes on the adult stage of the rice brown planthopper, Nilaparvata lugens. Entomol. Exp. Appl. 75:51–59; 1995.
Pusztai, A.; Ewan, S. W. B.; Grant, G.; Peumans, W. J. The relationship between survival and binding of plant lectins during small intestinal passage and their effectiveness as growth factors. Digestion 46:308–316; 1990.
Rao, K. V.; Rathore, K. S.; Hodges, T. K.; Fu, X.; Stoger, E.; Sudhakar, D.; Williams, S.; Christou, P.; Bharathi, M.; Brown, D. P.; Powell, K. S.; Spence, J.; Gatehouse, A. M.; Gatehouse, J. A. Expression of snowdrop lectin (GNA) in transgenic rice plants confers resistance to rice brown planthopper. Plant J. 15:469–477; 1998.
Schocher, R. J.; Shillito, R. D.; Saul, M. W.; Paszkowski, J.; Potrykus, I. Cotransformation of unlinked foreign genes into plants by direct gene transfer. Bio/Technology 4:1093–1096; 1986.
Shi, Y.; Wang, M. B.; Powell, K. S.; Van Damme, E.; Hilder, V. A.; Gatehouse, A. M. R.; Boulter, D.; Gatehouse, J. A. Use of the rice sucrose synthase-1 promoter to direct phloem-specific expression of β-glucuronidase and snowdrop lectin genes in transgenic tobacco plants. J. Exp. Bot. 45:623–631; 1994.
Sogawa, K.; Pathak, M. D. Mechanisms of brown planthopper resistance in the Mudgo variety of rice. Appl. Entomol. Zool. 5:145–158; 1970.
Song, W. Y.; Wang, G. L.; Chen, L. L.; Kim, H. S.; Pi, L. Y.; Holsten, T.; Gardner, J.; Wang, B.; Zhai, W. X.; Zhu, L. H.; Fauquet, C.; Ronald, P. A receptor kinase-like protein encoded by the rice disease resistance gene, Xa21. Science 270:1804–1806; 1995.
Sudhakar, D.; Fu, X.; Stoger, E.; Williams, S.; Spence, J.; Brown, D. P.; Bharathi, M.; Gatehouse, J. A.; Christou, P. Expression and immunolocalization of the snowdrop lectin insecticidal protein GNA, in transgenic rice plants. Transgenic Res. 7:371–378; 1998.
Tang, K., Tinjuangjun, P.; Xu, Y. Sun, X.; Gatehouse, J. A.; Ronald, P. C.: Qi, H.; Lu, X.; Christou, P.; Kohli, A.: Particle bombardmentmediated co-transformation of elite Chinese rice cultivars with genes conferring resistance to bacterial blight and sap sucking insect pests. Planta 208:552–563; 1999.
Wakita, Y.; Otani, M.; Iba, K.; Shimada, T. Co-integration, co-expression and co-segregation of an unlinked selectable marker gene and NtFAD3 gene in transgenic rice plants produced by particle bombardment. Genes Gen. Sys. 73:219–226; 1998.
Wang, M. B.; Boulter, D.; Gatehouse, J. A. A complete sequence of the rice sucrose synthase-1 (Rss1) gene. Plant Mol. Biol. 19:881–885; 1992.
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Tang, K., Hu, Q., Sun, X. et al. Development of transgenic rice pure lines with enhanced resistance to rice brown planthopper. In Vitro Cell.Dev.Biol.-Plant 37, 334–340 (2001). https://doi.org/10.1007/s11627-001-0060-8
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DOI: https://doi.org/10.1007/s11627-001-0060-8