Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/89334
標題: 構築洋桔梗壞疽病毒之轉基因抗性
Construction of Transgenic Resistance against Lisianthus necrosis virus
作者: Yu-Wei Lin
林郁瑋
關鍵字: 洋桔梗壞疽病毒
轉基因抗性
反向重複序列構築
洋桔梗
Lisianthus necrosis virus
transgenic resistance
inverted-repeat construct
lisianthus
引用: 何婉清。2003。萎凋病。植物保護圖鑑系列13-洋桔梗保護: 55-60。行政院農業委員會動植物防疫檢疫局。 沈原民、趙佳鴻、王妃蟬、劉興隆。2010。洋桔梗露菌病之發生及預防策略臺中區農業專訊 70: 16-19。 李敏郎。2003。莖枯病。植物保護圖鑑系列13-洋桔梗保護: 34-36。行政院農業委員會動植物防疫檢疫局。 孫文章、王瑞章、陳俊仁、胡文若。2005。外銷洋桔梗栽培技術。台南區農業改良場技術專刊。行政院農業委員會台南區農業改良場。6頁。 陳慶忠、陳煜焜、柯文華、徐惠迪。2002。康乃馨上洋桔梗壞疽病毒之分離與鑑 定。植物病理學會刊 11: 137-146。 陳慶忠、柯文華、陳煜焜。1998。洋桔梗上胡瓜嵌紋病毒鑑定及傳播試驗。臺中區農業改良場研究彙報 60: 1-18。 陳慶忠、柯文華、曹淑麗、趙佳鴻。2001。洋桔梗上蠶豆萎凋病毒之分離與鑑定。臺中區農業改良場研究彙報 70: 51-63。 陳彥樺、蔡宛育。2012。洋桔梗。臺灣中部地區外銷作物產業特刊 112: 110-118。行政院農業委員會臺中區農業改良場。 楊秀珠、謝廷芳。1998。洋桔梗露菌病的發生與藥劑防治植物保護學會會刊 40: 37-48。 劉興隆、曾國欽。2003。洋桔梗細菌性萎凋病。植物保護圖鑑系列13-洋桔 梗保護: 65-69。行政院農業委員會動植物防疫檢疫局。 蔡宛育、陳彥樺。2012。國內洋桔梗秋冬季主要栽培品種介紹。臺中區農業改良場特刊 110: 13-17。行政院農業委員會臺中區農業改良場。 蔡宛育、陳彥樺、陳姿翰。2012。洋桔梗國內產銷現況。臺中區農業改良場特刊 110: 5-11。行政院農業委員會臺中區農業改良場。 Abel, P. P., Nelson, R. S., De, B., Hoffmann, N., Rogers, S. G., Fraley, R. T., and Beachy, R. N. 1986. Delay of disease development in transgenic plants that express the Tobacco mosaic virus coat protein gene. Science 232: 738-743. An, Y. Q., McDowell, J. M., Huang, S., McKinney, E. C., Chambliss, S., and Meagher, R. B. 1996. Strong, constitutive expression of Arabidopsis ACT2/ACT8 actin subclass in vegetative tissues. Plant J. 10: 107-121. Audy, P., Palukaitis, P., Slack, S. A., and Zaitlin, M., 1994. Replicase-mediated resistance to Potato virus Y in transgenic plants. Mol. Plant-Microbe Interact. 7: 15-22. Baulcombe, D. C. 1996. Mechanisms of pathogen-derived resistance to viruses in transgenic plants. Plant Cell 8: 1833-1844. Brodersen, P., and Voinnet, O. 2006. The diversity of RNA silencing pathways in plants. Trends Genet. 22: 268-280. Chandler, S. F., and Sanchez, C. 2012. Genetic modification; the development of transgenic ornamental plant varieties. Plant Biotechnol. J. 10: 891-903. Chang, C. A., and H. T. Tsai. 1993. Isolation of Bean yellow mosaic virus from lisianthus developing foliar mosaic and flower breaking symptoms. Plant Pathol. Bull. 2: 250-251. Chang, C. H. 2003. Complete Genome Sequence and Genetic Organization of Lisianthus Necrosis Virus. Master thesis. Department of Plant Pathology, National Chung Hsing University, Taichung, Taiwan. 70pp. Chang, Y. S. 2007. Identification and Characterization of a Calla Lily-Infecting Isolate of Lisianthus Necrosis Virus. Master thesis. Department of Plant Pathology, National Chung Hsing University, Taichung, Taiwan. 53pp. Chang, C., Chen, Y.-C., Hsu, Y.-H., Wu, J.-T., Hu, C.-C., Chang, W.-C. and Lin, N.-S. 2005. Transgenic resistance to Cymbidium mosaic virus in Dendrobium expressing the viral capsid protein gene. Transgenic Res. 14: 41-46. Chao, C. H., Chen, C. C., Chang, C. A., and Chen, C. C. 2000. Identification of a Turnip mosaic virus isolate causing systemic yellow spotting on lisianthus. Plant Pathol. Bull. 9: 115-122. Chen, C. C., Chen, Y. K., and Hsu, H. T. 2000. Characterization of a virus infecting lisianthus. Plant Dis. 84: 506-509. Chen, C. C., Tsao, S. L., and Hsu, H. T. 2001. Diagnosis of Lisianthus necrosis virus infection by light and electron microscopy and serological assays. Plant Pathol. Bull. 10: 105-114. Chen, C. C., and Hsu, H. T. 2002. Occurrence of a severe strain of Lisianthus necrosis virus in imported carnation seedlings in Taiwan. Plant Dis. 86: 444. Chen, Y. K., Jan, F. J., Chen, C. C., and Hsu, H. T. 2006. A new natural host of Lisianthus necrosis virus in Taiwan. Plant Dis. 90: 1112. Chen, Y. K., Lohuis, D., Goldbach, R., and Prins, M. 2004. High frequency induction of RNA-mediated resistance against Cucumber mosaic virus using inverted repeat constructs. Mol. Breed. 14: 215-226. Chen, Y. K., Chang, Y. S., and Chen, C. C. 2011. Identification of Carnation mottle virus from lisianthus plants in Taiwan. Plant Dis. 95: 1036. Chen, Y. T., Fang, Q. S., Chiang, C. H., Yeh, S. D., Wu, H. W., and Yu, T. A. 2010. Transgenic Eustoma grandiflorum expressing the bar gene are resistant to the herbicide BastaR. Plant Cell Tiss. Organ. Cult. 102: 347-356. Cheng, Y. H., Chen, C. C., and Chang, C. A. 2005. Whitefly-transmitted geminiviruses in ornamental plants and their control strategies in Taiwan. Page 95-105 in Proceedings of the International Seminar on Whitefly Management and Their Control Strategy. Food and Fertilizer Tech. Cent. and Agric. Res. Inst. Pub. Taichung, Taiwan. Cheng, Y. H., Chen, C. C., Liao,J. Y., Deng, T. C., and Chang, C. A. 2009. Isolation and identification of Pepper veinal mottle virus on lisianthus. J. Taiwan Agric. Res. 58: 196-207. Christie, S. R., Purcifull, D. E., Crawford, W. E., and Ahmed, N. A. 1985. Electron microscopy of negatively stained clarified viral concentrates obtained from small tissue samples with appendices on negative staining techniques. Fla. Agr. Sta. Bull. No. 872. 45 pp. Chu, M., Park, J.-W., and Scholthof, H. B. 1999. Separate regions on the Tomato bushy stunt virus p22 protein mediate cell-to-cell movement versus elicitation of effective resistance responses. Mol. Plant-Microbe Interact. 12: 285-292. Chu, M., Desvoyes, B., Turina, M., Noad, R. and Scholthof, K. B. G. 2000. Genetic dissection of Tomato bushy stunt virus p19- protein-mediated host-dependent symptom induction and systemic invasion. Virology 266: 79-87. Cohen, J., Gera, A., Ecker, R., Ben, J. R,. Perlsman, M., Gokkes, M., Lachman, O., and Antignus, Y. 1995. Lisianthus leaf curl, a new disease of lisianthus caused by Tomato yellow leaf curl virus. Plant Dis. 79: 416-420. Cohen, J., Lapidot, M., Loebenstein, G., and Gera, A. 2001. First report of Sweet potato sunken vein virus occurring in lisianthus. Plant Dis. 85: 697. de Freitas, J. C., Kitajima, E. W., and Rezende, J. A. M. 1996. First report of Tobacco streak virus on lisianthus in Brazil. Plant Dis. 80:1080. Di Nicola-Negri, E., Brunetti, A., Tavazza, M., and Ilardi, V. 2005. Hairpin RNA-mediated silencing of Plum pox virus P1 and HC-Pro genes for efficient and predictable resistance to the virus. Transgenic Res. 14: 989-994. El-Hamalawi, Z. A. and M. E. Stanghellini. 2005. Disease development on lisianthus following aerial transmission of Fusarium avenaceum by adult shore flies, fungus gnats, and moth flies. Plant Dis. 89: 619-623. Fauquet, C. M., Mayo, M. A., Maniloff, J., Desselberger, U., and Ball, L. A. 2005. Virus Taxonomy: Eighth Report of the International Committee on Taxonomy of Viruses. Elsevier Academic Press, USA. 1162pp. Gera, A. G. and Cohen, J. 1990. The natural occurrence of Bean yellow mosaic, Cucumber mosaic and Tobacco mosaic viruses in lisianthus in Israel. Plant Pathol. 39: 561-564. Goldbach, R., Bucher, E., and Prins, M. 2003. Resistance mechanisms to plant viruses: an overview. Virus Res. 92: 207-212. Handa, T. 1992. Regeneration and characterization of prairie gentian (Eustoma grandiforum) plant transformed by Agrobacterium rhizogenes. Plant Tiss. Cult. Lett. 9: 10-14. Harlow, E., and Lane, D. 1988. Antibodies-A Laboratory Manual. Cold Spring Harbor Press. 726pp. Hearne, P. Q., Knorr, D.A., Hillman, B. and Morris, T.J. 1990. The complete genome structure and synthesis of infectious RNA from clones of Tomato bushy stunt virus. Virology 177: 141-151. Horsch, R. B., Fry, J. E., Hoffmann, N. L., Eichholtz, D., Rogers, S. G., and Fraley, R. T. 1985. A simple and general method for transferring genes into plants. Science 227: 1229-1231. Iwaki, M., Hanada, K., Maria, E. R. A., and Onogi, S. 1987. Lisianthus necrosis virus, a new necrovirus from Eustoma russellianum. Phytopathology 77: 867-870. Iwaki, M., Maria, E.R.A., Hanada, K., Onogi, S. and Zenbayashi, R. 1985. Three viruses occurred in lisianthus plants. Ann. Phytopathol. Soc. Japan 52: 355. Jan, F. J., Zheng, Y. X., Chao, C. H., Ko, W. F., Chang, C. C., and Chen, C. C. 2003. Identification of a tobamovirus causing yellow mottle and stunting symptoms on lisianthus in Taiwan. Plant Pathol. Bull. 12:122–132. Kawaike, N., Sato, T., and Kawada,Y. 1998. Occurrence and control of the brown spot of prairie gentian, Eustoma grandiflorum (Raf.) Shinn. caused by Stemphylium lycopersici in Kochi Prefecture. Proc. Assoc. Plant Prot. Shikoku 33: 43-48. King, A. M. Q., Adams, M. J., Carstens, E. B., and Lefkowitz, E. J. 2012. Virus Taxonomy – Classification and Nomenclature of Viruses. Ninth Report of the International Committee on Taxonomy of Viruses. Elsevier Academic Press, Amsterdam. 1327pp. Koenig, R. 1981. Indirect ELISA methods for the broad specificity detection of plant viruses. J. Gen. Virol. 55: 53-62. Kritzman, A., Beckelman, H., Alexandrov, S., Cohen, J., Lampel, M., Zeidan, M., Raccah, B., and Gera, A. 2000. Lisianthus leaf necrosis: a new disease of lisianthus caused by Iris yellow spot virus. Plant Dis. 84: 1185-1189. Ledger, S. E., Deroles, S. C., Manson, D. G., Bradley, J. M., and Given, N. K. 1997. Transformation of lisianthus (Eustoma grandiflorum). Plant Cell Rep. 16: 853-858. Liao, L. J., Pan, I. C., Chan, Y. L., Hsu, Y. H., Chen, W. H. and Chan, M. T. 2004. Transgene silencing in Phalaenopsis expressing the coat protein of Cymbidium mosaic virus is a manifestation of RNA-mediated resistance. Mol. Breed. 13: 229-242. Lisa, V. and Dellavalle, G. 1987. Bean yellow mosaic virus in Lisianthus russellianus. Plant Pathol. 36: 214-215. Lisa, V., Varia, A. M., d'Aquilio, M., Dellavalle, G., Masenga, G., Milne, R. G., and Bocardo, G. 1994. Characterization of an Ilarvirus from lisianthus (Eustoma grandiflorum). Acta. Hortic. 377: 81-89. Lindbo, J. A., and Dougherty, W. G. 1992. Pathogen-derived resistance to a potyvirus: immune and resistance phenotypes in transgenic tobacco expressing altered forms of a potyvirus coat protein nucleotide sequence. Mol. Plant-Microbe Interact. 5: 144-153. Lindbo, J. A., Silva-Rosales, L., Proebsting, W. M., and Dougherty, W. G. 1993. Induction of a highly specific antiviral state in transgenic plants: implications for regulation of gene expression and virus resistance. Plant Cell 5: 1749-1759. McGovern, R. J., Polston, J. E., and Harbaugh, B. K. 1997. Detection of a severe isolate of Impatiens necrotic spot virus infecting lisianthus in Folrida. Plant Dis. 81:1334. McGovern, R. J., Bouzar, H., and Harbough, B. K. 2000. Stem blight of Eustoma grandiflorum caused by Sclerotium rolfsii. Plant Dis. 84: 490. Makkouk, K. M., Koenig, R., and Lessemann, D. E. 1981. Characterization of a Tombusvirus isolated from eggplant. Phytopathology 71: 572-577. Missiou, A., Kalantidis, K., Boutla, A., Tzortzakaki, S., Tabler, M., and Tsagris, M. 2004. Generation of transgenic potato plants highly resistant to potato virus Y (PVY) through RNA silencing. Mol. Breed. 14: 185-197. Mueller, E., Gilbert, J., Davenport, G., Brigneti, G., and Baulcombe, D. C. 1995. Homology-dependent resistance: transgenic virus resistance in plants related to homology-dependent gene silencing. Plant J. 7: 1001-1013. Nordlee, J. A., Taylor, S. L., Townsend, J. A., Thomsa, L. A., and Bush, R. K. 1996. Identification of a Brazil-nut allergen in transgenic soybeans. New Engl. J. Med. 334: 688-692. Niu, Q. W., Lin, S. S., Reyes, J. L., Chen, K. C., Wu, H. W., Yeh, S. D., and Chua, N. H. 2006. Expression of artificial microRNAs in transgenic Arabidopsis thaliana confers virus resistance. Nat. biotechnol. 24: 1420-1428. Ono, E., Fukuchi-Mizutani, M., Nakamura, N., Fukui, Y., Yonekura-Sakakibara, K., Yamaguchi, M., Nakayama, T., Tanaka, T., Kusumi, T. and Tanaka, Y. 2006. Yellow flowers generated by expression of the aurone biosynthetic pathway. Proc. Natl. Acad. Sci. 103: 11075-11080. Pang, S. Z., Jan, F. J., and Gonsalves, D. 1997. Nontarget DNA sequences reduce the transgene length necessary for RNA-mediated tospovirus resistance in transgenic plants. Proc. Natl. Acad. Sci. 94: 8261-8266. Prins, M., de Oliveira Resende, R., Anker, C., Van Schepen, A., Haan, P. D., and Goldbach, R. 1996. Engineered RNA-mediated resistance to tomato spotted wilt virus is sequence specific. Mol. Plant-Microbe Interact. 9: 416-418. Provvidenti, R. 1985. Two newly recognized hosts of Cucumber mosaic virus: Eustoma grandiflorum and Peristrophe angustiofolia. Plant Dis. 69: 524. Reynolds, E. S. 1963. The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J. Cell. Biol. 17: 208-212. Rochon, D., Lommel, S. A., Martelli, G. P., Rubino, L., and Russo, M. 2012. Tombusviridae. Pages 1111-1138. in King, A. M. Q., Adams, M. J., Carstens, E. B., and Lefkowitz, E. J. (Eds.) Virus Taxonomy – Classification and Nomenclature of Viruses. Ninth Report of the International Committee on Taxonomy of Viruses. Elsevier Academic Press, Amsterdam. 1327pp. Russo, M., Vovlas, C., Rubino, L., Grieco, F., and Martelli, G. P. 2002. Molecular characterization of a tombusvirus isolated from diseased pear trees in southern Italy. J. Plant Pathol. 84: 1961-1966. Sanford, J. C., and Johnston, S. A. 1985. The concept of parasite-derived resistance -Deriving resistance genes from the parasite's own genome. J. Theoret. Biol. 113: 395-405. Saxena, D., Flores, S., and Stotzky, G. 1999. Insecticidal toxin in root exudates from Bt corn. Nature 402: 480. Schochow, M., Tjosvold, S. A., and Ploeg, A. T. 2004. Host status of lisianthus 'Mariachi Lime Green'for three species of root-knot nematodes. Hort. Sci. 39: 120-123. Scholthof, H. B., Scholthof, K. B. G., Kikkert, M. and Jackson, A. O. 1995. Tomato bushy stunt virus spread is regulated by two nested genes that function in cell-to-cell movement and host dependent systemic invasion. Virology 213: 425-438. Schouten, A., Roosien, J., de Boer, J. M., Wilmink, A., Rosso, M. N., Bosch, D., Stiekema, W. J., Gommers, F. J., Bakker, J., and Schots, A. 1997. Improving scFv antibody expression levels in the plant cytosol. FEBS Letters 415: 235-241. Semeria, L., Vaira, A. M., Accotto, G. P., and Allavena, A. 1995. Genetic transformation of Eustoma grandiforum Griseb. by microinjectile bombardment. Euphytica 85: 125-130. Semeria, L., Ruffoni, B., Rabaglio, M., Genga, A., Vaira, A. M., Accotto, G. P., and Allavena, A. 1996. Genetic transformation of Eustoma grandiflorum by Agrobacterium tumefaciens. . Plant Cell Tiss. Org. Cult. 47: 67-72. Shen, Y. M., C. H. Chao, F. C. Wang, H. L. Liu and T. C. Huang. 2012. First report of stem and leaf blight caused by Sclerotinia sclerotiorum on eustoma in Taiwan. Plant Dis. 96:910. Shpialter, L., D. Rav David, I. Dori, U. Yermiahu, S. Pivonia, R. Levite and Y. Elad. 2009. Cultural methods and environmental conditions affecting gray mold and its management in lisianthus. Phytopathology 99: 557-570. Singh, O. V., Ghai, S., Paul, D., and Jain, R. K. 2006. Genetically modified crops: success, safety assessment, and public concern. Appl. Microbiol. Biotechnol. 71: 598-607. Spitzer-Rimon, B., Marheva, E., Barkal, O., Marton, I., Edelbaum, O., Masci, T., Naveen-Kumar, P., Shklamann, E., Ovadis, M. and Vainstein, A. 2010. EOBII, a gene encoding a flower-specific regulator of phenylpropanoid volatiles' biosynthesis in petunia. Plant Cell 22: 1961-1976. Smith, N. A., Singh, S. P., Wang, M. B., Stoutjesdijk, P. A., Green, A. G., and Waterhouse, P. M. 2000. Total silencing by intron spliced hairpin RNAs. Nature 407: 319-320. Stam, M., Mol, J. N., and Kooter, J. M. 1997. The silence of genes in transgenic plants. Annals Bot. 79: 3-12. Stubbs, L.L. 1947. First description of wilting disease in broad bean. J. Dep. Agric. Vict. 46: 323. Suzuki, M., M. Togawa and N. Sato. 2003. Control effects of soil-borne diseases of Russell prairie gentian (Eustoma grandiflorum) by watering after steam sterilization. Bull. Shizuoka Agr. Exp. Sta. 48: 29-33. Tougou, M., Furutani, N., Yamagishi, N., Shizukawa, Y., Takahata, Y., and Hidaka, S. 2006. Development of resistant transgenic soybeans with inverted repeat-coat protein genes of soybean dwarf virus. Plant Cell Rep. 25: 1213-1218. van Engelen, F. A., Schouten, A., Molthoff, J. W., Roosien, J., Salinas, J., Dirkse, W. G., Schots, A., Bakker, J., Gommers, F. J., Jongsma, M. A., Bosch, D., and Stiekema, W. J. 1994. Coordinate expression of antibody subunit genes yields high levels of functional antibodies in roots of transgenic tobacco. Plant Mol. Biol. 26: 1701-1710. Vrind, T. A. 2005. The botrytis problem in figures. Acta Hortic. 669: 99-102. Voinnet, O. 2005. Induction and suppression of RNA silencing: insights from viral infections. Nature Rev. Genet. 6: 206-220. Wang, M. B., Abbott, D. C., and Waterhouse, P. M. 2000. A single copy of a virus‐derived transgene encoding hairpin RNA gives immunity to barley yellow dwarf virus. Mol. Plant Pathol. 1: 347-356. Wang, M. B., Masuta, C., Smith, N. A., and Shimura, H. 2012. RNA silencing and plant viral diseases. Mol. Plant-Microbe Interact. 25: 1275-1285. Waterhouse, P. M., Graham, M. W., and Wang, M. B. 1998. Virus resistance and gene silencing in plants can be induced by simultaneous expression of sense and antisense RNA. Proc. Nat. Acad. Sci. 95: 13959-13964. Warner, R. 2011. Genetic approaches to improve cold tolerance of petunia. Floricult. Int. 15-16. Wegulo, S. N. and Vilchez, M. 2007. Evaluation of lisianthus cultivars for resistance to Botrytis cinerea. Plant Dis. 91: 997-1001. Yang, C. F., Chen, K. C., Cheng, Y. H., Raja, J. A., Huang, Y. L., Chien, W. C., and Yeh, S. D. 2014. Generation of Marker-free Transgenic Plants Concurrently Resistant to a DNA Geminivirus and a RNA Tospovirus. Sci. Rep. 4: 5717. Zaccai, M., Lewinsohn, E., and Pichersky, E. 2001. Modifying lisianthus traits by genetic engineering. Acta Hort. 552: 137-142. Zhang, Z. Y., Wang, Y. G., Shen, X. J., Li, L., Zhou, S. F., Li, W. C., and Fu, F. L. 2013. RNA interference-mediated resistance to maize dwarf mosaic virus. Plant Cell Tiss. Org. Cult. 113: 571-578.
摘要: 洋桔梗 (Eustoma russellianum (Don.) Griseb) 為重要的切花類觀賞花卉,在洋桔梗的栽培過程中常受到病毒病害的感染,其中以洋桔梗壞疽病毒 (Lisianthus necrosis virus, LNV) 所造成的危害最具威脅性。為了使植物具備病毒抗性,目前已有傳統的抗病育種,以及建構轉基因抗性等方法。但抗病育種所需時間較長,相對而言轉基因抗性較為省時又能表現穩定專一的抗性,因此成為建構病毒抗性之主流。本研究選擇使用後轉錄基因靜默機制 (post-transcriptional gene silencing, PTGS)誘導洋桔梗產生病毒抗性。根據前人研究,不論將正向或反向的病毒序列導入植物體內皆能誘導siRNA產生進而引發PTGS,其中又以反向重複序列的構築方式能提供最佳的抗病效率。針對LNV的基因體中RdRP、CP、MP等基因設計了四個目標區域,以反向重複序列的方式構築於二元載體pBIN (pLNVRD5-IR、pLNVRD3-IR、pLNV3'CP-IR、pLNVMP-IR),利用農桿菌轉殖法 (Agrobacterium tumefaciens-mediated transformation, ATMT)將其導入模式植物圓葉菸草 (Nicotiana benthamiana)及洋桔梗 (Eustoma russellianum)中。菸草R0轉殖株的結果LNV RdRP基因的前半 (5'端)所構築的反向重複序列 (pLNVRD5-IR)所提供的轉基因抗性為最高,可達到80.1%,其餘三種反向重複序列構築- RdRP基因的後半 (3'端) (pLNVRD3-IR)、外鞘蛋白基因的後半 (3'端) (pLNV3'CP-IR)、移動蛋白基因 (pLNVMP-IR)-亦可使轉殖株產生對LNV之抗性,抗病比率依序為32%、41.2%與12.5%。相同的構築已轉殖至洋桔梗,因轉殖株的生長速度較慢,目前仍在進行中,尚未進行抗病性的相關測試。
Lisianthus (Eustoma russellianum (Don.) Griseb) is an important ornamental crop in the cut flower industry worldwide. However, there are several viral diseases interfere lisianthus production in Taiwan, especially Lisianthus necrosis virus (LNV), which is probably the most devastating one to lisianthus. There were many approaches, including conventional resistance breeding and transgenic resistance, have been developed to build resistance in plants against viral diseases. Compared to the time-consuming conventional resistance breeding, transgenic approaches can provide more stable and specific resistance efficiency. In this study, in order to provide lisianthus with virus resistance, the effective strategy of post-transcriptional gene silencing (PTGS) was chosen. Based on previous researches, transgenic plants harboring viral sequences, both in sense or in anti-sense orientations, can trigger the formation of siRNA and thus lead to PTGS. These have been demonstrated successfully in several crops. Among these, the approach of using inverted-repeat (IR) constructs has been shown to have better efficacy. The cDNA fragments derived from LNV genes of RNA-dependent RNA polymerase (RdRp), coat protein (CP) and movement protein (MP) were separately cloned into the binary vector pBIN in the form of IR and already introduced into Nicotiana benthamiana and Eustoma russellianum by Agrobacterium tumefaciens-mediated transformation (ATMT) to generate transgenic tobacco and lisianthus potentially resistant to LNV. All four IR constructs, i.e. pLNVRD5-IR, pLNVRD3-IR, pLNV3'CP-IR, and pLNVMP-IR, conferred transgenic resistance to tobacco plants and the pLNVRD5-IR conferred the best efficiency among the four constructs. The resistance ratio in R0 tobacco of constructs pLNVRD5-IR, pLNVRD3-IR, pLNV3'CP-IR and pLNVMP-IR are 80.1%, 32%, 41.2% and 12.5%, respectively. The transformation and regeneration of lisianthus are now under processing. However, due to the limitation of growth rate of tissue-cultured lisianthus, the transgenic resistance of these transformants have not been analyzed yet.
URI: http://hdl.handle.net/11455/89334
文章公開時間: 10000-01-01
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