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標題: 抗胡瓜嵌紋、胡瓜綠斑嵌紋與西瓜嵌紋病毒轉基因西瓜之研發
Development of transgenic watermelon plants with multiple resistance to Cucumber mosaic-、Cucumber green mottle mosaic- and Watermelon mosaic viruses
作者: 何琇銀
Ho, Hsiu-Yin
關鍵字: transgenic watermelon;轉基因西瓜;Cucumber mosaic virus;Cucumber green mottle mosaic virus;Watermelon mosaic virus;胡瓜嵌紋病毒;胡瓜綠斑嵌紋病毒;西瓜嵌紋病毒
出版社: 農藝學系所
引用: 沈百奎、鄧汀欽、余志儒、林俊義。2002。西瓜栽培管理。行政院農業委員會農業試驗所特刊第103號。台中。44pp。 李青梅。2005。西瓜銀斑病毒核鞘蛋白與矮南瓜黃化嵌紋病毒及木瓜輪點病毒西瓜系統鞘蛋白轉基因西瓜之構築。大葉大學分子生物科技學系碩士論文。台中。 林學詩。1995。西瓜及洋香瓜產業與研究。臺灣蔬菜產業改進研討會專集: 7-20。台中區農業改良場特刊第37號。台中。369pp。 施傑仁。2002。利用基因沉寂機制發展抗多種病毒病害之轉基因茄科植物。中興大學植物病理學系碩士論文。台中。 林靜宜、詹富智。2005。無篩選標示基因轉基因植物之構築及其最新發展。植物病理學會刊14: 159-176。 張治國。2004。台灣西瓜產銷概況與展望。農政與農情147: 61-67。 Akashi, K., Morikawa, K., and Yokota, A. 2005. Agrobacterium-mediated transformation system for the drought and excess light stress-tolerant wild watermelon (Citrullus lanatus). Plant Biotechnol. 22: 13-18. Arce-Ochoa, J. P., Dainello, F., Pike, L. M., and Drews, D. 1995. Field performance comparison of two transgenic summer squash hybrids to their parental hybrid line. HortScience 30: 492-493. Baulcombe, D. C. 1996. RNA as a target and an initiator of post-transcriptional gene silencing in transgenic plants. Plant Mol. Biol. 32: 79-88. Baulcombe, D. C. 2004. RNA silencing in plants. Nature 431: 356-363. Bennett, P. M., Livesey, C. T., Nathwani, D., Reeves, D. S., Saunder, J. R., and Wise, R. 2004. An assessment of the risks associated with the use of antibiotic resistance genes in genetically modified plants: report of the Working Party of the British Society for Antimicrobial Chemotherapy. J. Antimicrob. Chemother. 53: 418-431. Chang, Y. M., Hsaio, C. H., Yang, W. Z., Hseu, S. H., Chao, Y. J., and Huang, C. H. 1987. The occurrence and distribution of five cucurbit viruses on melon and watermelon in Taiwan. J. Agri. Res. China 36: 389-397. Chee, P. P., and Slightom, J. L. 1991. Transfer and expression of cucumber mosaic virus coat protein gene in the genome of Cucumis sativus. J. Am. Soc. Hortic. Sci. 116: 1098-1102. Chen, C. C., Ho, H. M., Chang, T. F., Chao, C. H., and Yeh, S. D. 1995. Characterization of a tospovirus-like virus isolated from wax gourd. Plant Prot. Bull. 37: 117-131. Chen, T. C., Lu, Y. Y., Cheng, Y. H., Chang, C. A., and Yeh, S. D. 2006. Characterization of a tospovirus isolated from central Taiwan as Melon yellow spot virus. Bull. Plant Pathol. 15: 307-308. Chen, W. S., Chiu, C. C., Liu, H. Y., Lee, T. L., Cheng, J. T., Lin, C. C., Wu, Y. J., and Chang, H. Y. 1998. Gene transfer via pollen-tube pathway for anti-fusarium wilt in watermelon. Biochem. Mol. Biol. Int. 46: 1201-1209. Choi, G. S. 2001. Occurrence of two Tobamovirus diseases on cucurbits and control measures in Korea. Plant Pathol. J. 17: 243-248. Choi, P. S., Soh, W. Y., Kim, Y. S., Yoo, O. J., and Liu, J. R. 1994. Genetic transformation and plant regeneration of watermelon using Agrobacterium tumefaciens. Plant Cell Rep. 13: 344-348. Clough, G. H., and Hamm, P. B. 1995. Coat protein transgenic resistance to watermelon mosaic and zucchini yellows mosaic virus in squash and cantaloupe. Plant Dis. 79: 1107-1109. Compton, M. E., and Gray, D. J. 1993. Shoot organogenesis and plant regeneration from cotyledons of diploid, triploid, and tetraploid watermelon. J. Am. Soc. Hortic. Sci. 118: 151-157. Compton, M. E. 1999. Dark pretreatment improves adventitious shoot organogenesis from cotyledons of diploid watermelon. Plant Cell Tissue Organ Cult. 58: 185-188. Compton, M. E. 2000. Interaction between explant size and cultivar impacts shoot organogenic competence of watermelon cotyledons. HortScience 35: 749-750. Compton, M. E., Gray, D. J., and Gaba, V. P. 2004. Use of tissue culture and biotechnology for the genetic improvement of watermelon. Plant Cell Tissue Organ Cult. 77: 231-243. Dabauza, M., Bordas, M., Salvador, A., Roig, L. A., and Moreno, V. 1997. Plant regeneration and Agrobacterium-mediated transformation of cotyledon explants of Citrullus colocynthis (L.) Schrad. Plant Cell Rep. 16: 888-892. Dale, P. J., Clarke, B., and Fontes, E. M. G. 2002. Potential for the environmental impact of transgenic crops. Nat. Biothchnol. 20: 567-574. de Haan, P., Gielen, J. J., Prins, M., Wijkamp, I. G., van Schepen, A., Peters, D., van Grinsven, M. Q., and Goldbach, R. 1992. Characterization of RNA-mediated resistance to tomato spotted wilt virus in transgenic tabacco plants. Bio/Technology 10: 1133-1137. Deng, T. C., Tsai, C. H., Chen, Y. F., and Chang, C. A. 1997. First report of cucurbit aphid-borne yellows luteovirus in Taiwan. Plant Prot. Bull. 39: 395-396. Desbiez, C., and Lecoq, H. 2004. The nucleotide sequence of Watermelon mosaic virus (WMV, Potyvirus) reveals interspecific recombination between two related potyviruses in the 5'' part of the genome. Arch.Virol. 149: 1619-1632. Dong, J. Z., and Ji, S. R. 1991. High efficiency plant regeneration from cotyledons of watermelon (Citrullus vulgaris Schrad). Plant Cell Rep. 9: 559-562. Doolittle, S. P. 1916. A new infectious mosaic disease of cucumber. Phytopathology 6: 145-147. Ellul, P., Rios, G., Atares, A., Roig, L. A., Serrano, R., and Moreno, V. 2003. The expression of the Saccharomyces cerevisiae HAL1 gene increases salt tolerance in transgenic watermelon [Citrullus lanatus (Thunb.) Matsun. & Nakai.]. Theor. Appl. Genet. 107: 462-469. Feinberg, A. P., and Vogelstein, B. 1983. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal. Biochem. 132: 6-13. Francki, R.I.B., Mossop, D.W., and Hatta, T. 1979. Cucumber mosaic virus. CMI/AAB Descriptions of Plant Viruses, No. 213. Fuchs, M., and Gonsalves, D. 1995. Resistance of transgenic hybrid squash ZW-20 expressing the coat protein genes of zucchini yellow mosaic virus and watermelon mosaic virus 2 to mixed infections by both potyviruses. Bio/Technology 13: 1466-1473. Fuchs, M., McFerson, J. R., Tricoli, D. M., Russell, M. J., Deng, R. Z., Boeshore, M. L., Reynolds, J. F., Russell, P. F., Quemada, H. D., and Gonsalves, D. 1997. Cantaloupe line CZW-30 containing coat protein genes of cucumber mosaic virus, zucchini yellow mosaic virus, and watermelon mosaic virus-2 is resistant to these three viruses in the field. Mol. Breed. 3: 279-290. Fuchs, M., Tricoli, D. M., Carney, K. J., Schesser, M., McFerson, J. R., and Gonsalves, D. 1998. Comparative virus resistance and fruit yield of transgenic squash with single and multiple coat protein genes. Plant Dis. 82:1350-1356. Fukuda, M., Meshi,T., Okada, Y., Otsuki, Y., and Takebe, I. 1981. Correlation between particle multiplicity and location on virion RNA of the assembly intiation site for viruses of the tobacco mosaic virus group. Proc. Natl. Acad. Sci. USA 78: 423l-4235. Fulton, T. M. 1995. Microprep protocol for extraction of DNA from tomato and other herbaceous plants. Plant Mol. Biol. Rep. 13: 207-209. Goldbach, R., Bucher, E., and Prins, M. 2003. Resistance mechanisms to plant viruses: an overview. Virus Res. 92: 207-212. Gonsalves, D., Chee, P., Provvidenti, R., Seem, R., and Slightom, J. 1992. Comparison of coat protein-mediated and genetically derived resistance in cucumber to infection by cucumber mosaic virus under field conditions with natural challenge inoculations by vectors. Bio/Technology 10: 1562-1570. Grumet, R. 1994. Development of virus resistant plants via genetic engineering. Plant Breed. Rev. 12: 47-49. Hseu, S. H., Huang, C. H., Chang, C. A., Yang, W. Z., Chang, Y. M., and Hsiao, C. H. 1987. The occurrence of five viruses in six cucurbit in Taiwan. Plant Prot. Bull. 29: 233-244. Hseu, S. H., Wang, H. L., and Huang, C. H. 1985. Identification of a zucchini yellow mosaic virus from Cucumis astivus. J. Agric. Res. China 34: 87-95. Hsu, H. T., Barzuna, L., Hsu, Y. H., Bliss,W., and Perry, K. L. 2000. Identification and subgrouping of Cucumber mosaic virus with mouse monoclonal antibodies. Phytopathology 90: 615-620. Hsu, H. T., Lawson, R. H., Yeh, S. D., and Chiu, R. J. 1996. Serological relationship of a high temperature-recovered tospovirus in the USA and watermelon silver mottle virus in Taiwan. Phytopathology 86: 843. Huang, C. H., Chang, L., and Tsai, J. H. 1993. The partial characterization of melon vein-banding mosaic virus, a newly recognized virus infecting cucurbits in Taiwan. Plant Pathol. 42: 100-107. Jagger, I. C. 1916. Experiments with the cucumber mosaic disease. Phytopathology 6: 149-151. Jan, F. J. 1998. Roles of nontarget DNA and viral gene length in influencing multi-virus resistance through homology-dependent gene silencing. Ph. D. Dissertation, Department of Plant Pathology, Cornell University. Jan, F. J., Fagoaga, C., Pang, S. Z., and Gonsalves, D. 2000a. A minimum length of N gene sequence in transgenic plants is required for RNA-mediated tospovirus resistance. J. Gen. Virol. 81: 235-242. Jan, F. J., Fagoaga, C., Pang, S. Z., and Gonsalves, D. 2000b. A single chimeric transgene derived from two distinct viruses confers multi-virus resistance in transgenic plants through homology-dependent gene silencing. J. Gen. Virol. 81: 2103-2109. Jan, F. J., Pang, S. Z., Tricoli, D. M., and Gonsalves, D. 2000c. Evidence that resistance in squash mosaic comovirus coat-protein-transgenic plants is affected by plant developmental stage and enhanced by combination of transgenes from different lines. J. Gen. Virol. 81: 2299-2306. Jan, F. J., Shih, J. R., Yeh, S. D., and Gonsalves, D. 2002. Development of transgenic plants resistant to multiple viruses via gene silencing. Page 14 in: Abstract book of XII International Congress of Virology, July 27-August 1, 2002, Paris, France. Jauhar, P. P. 2001. Genetic engineering and accelerated plant improvement: Opportunities and challenges. Plant Cell Tissue Organ Cult. 64: 87-91. Kamachi, S., Mochizuki, A., Nishiguchi, M., and Tabei, Y. 2007. Transgenic Nicotiana benthamiana plants resistant to cucumber green mottle mosaic virus based on RNA silencing. Plant Cell Rep. 26: 1283-1288. Lee, K. Y. 1996. Current occurrence and control of CGMMV Konjak disease. Plant Dis. Agric. 2: 38-39. Lomonossoff, G. P. 1995. Pathogen-derived resistance to plant viruses. Annu. Rev. Phytopathol. 33: 323-343. Miflin, B. 2000. Crop improvement in the 21st century. J. Exp. Bot. 51:1-8. 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. Namba, K., Pattanayek, R., and Stubbs, G. 1989. Visualization of protein-nucleic acid interactions in a virus. Refined structure of intact tobacco mosaic virus at 2.9 Å resolution by X-ray fiber diffraction. J. Mol. Biol. 208: 307-325. Napoli, C., Lemieux C., and Jorgenson, R. 1990. Introduction of a chimeric chalcone synthase gene into petunia results in reversible co-suppression of homologous genes in trans. Plant Cell 2: 279-289. Neves-Borges, A. C., Collares, W. M., Pontes, J. A., Breyne, P., Farinelli, L., and de Oliveira, D. E. 2001. Coat protein RNAs-mediated protection against Andean potato mottle virus in transgenic tobacco. Plant Sci. 160: 699-712. Nishibayashi, S., Hayakawa, T., Nakajima, T., Suzuki, M., and Kaneko, H. 1996. CMV protection in transgenic cucumber plants with an introduced CMV-O cp gene. Theor. Appl. Genet. 93: 672-678. 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. USA 94: 8261-8266. Pang, S. Z., Jan, F. J., Tricoli, D. M., Russell, P. F., Carney, K.J. Hu, J. S., Fuchs, M., Quemada, H. D., and Gonsalves, D. 2000. Resistance to squash mosaic comovirus in transgenic squash plants expressing its coat protein genes. Mol. Breed. 6: 87-93. Papayiannis, L. C., Ioannou, N., Boubourakas, I. N., Dovas, C. I., Katis, N. I., and Falk, B. W. 2005. Incidence of viruses infecting cucurbits in cyprus. J. Phytopathol. 153: 530-535. Park, S. M., Lee, J. S., Jegal, S., Jeon, B. Y., Jung, M., Park, Y. S., Han, S. L., Shin, Y. S., Her, N. H., Lee, J. H., Lee, M. Y., Ryu, K. H., Yang, S. G., and Harn, C. H. 2005. Transgenic watermelon rootstock resistant to CGMMV (cucumber green mottle mosaic virus) infection. Plant Cell Rep. 24: 350-356. Powell, P. A., Nelson, R. S., De, B., Hoffmann, N. L., Riogers, 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. Provvidenti, R. 1991. Inheritance of resistance to the Florida strain of zucchini yellow mosaic virus in watermelon. HortScience 26: 407-408. Provvidenti, R. 1996. Diseases caused by viruses. In: Zitter T.A., Hopkins D.L., Thomas C.E. (eds.). Compendium of Cucurbit Diseases, pp. 37-45. APS Press, St. Paul, Minn. Purcifull, D. E., Heibert, E., Petersen, M. A., Simone, G. W., Kucharek, T. A., Gooch, M. D., Crawford, W. E., Beckham, K. A., and De Sa, P. B. 1998. Partial characterization of a distinct potyvirus isolated from watermelon in Florida. Plant Dis. 82: 1386-1390. Rajagopalan, P. A., and Rafael, P. T. 2005. Improved cucumber transformation by a modified explant dissection and selection protocol. HortScience 40: 431-435. Roossinck, M. J. 2002. Evolutionary history of Cucumber mosaic Virus deduced by phylogenetic analyses. J. Virol. 76: 3382-3387. Sambrook, J., and Russell, D. W. 2001. Molecular Cloning: A Laboratory Manual, 3th ed. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory. Sanford, J. C., and Johnston, S. A. 1985. The concept of parasite-derived resistance - deriving resistance genes from the parasite''s own genome. J. Theor. Biol. 113: 395-405. Shukla, D. D., Ward, C. W., and Brunt, A. A. 1994. The Potyviridae. CAB International, Wallingford, UK. Smith, H. A., Swaney, S. L., Parks, T. D., Wernsman, E. A., and Dougherty, W. G. 1994. Transgenic plant virus resistance mediated by untranslatable sense RNAs: Expression, regulation, and fate of nonessential RNAs. Plant Cell 6: 1441-1453. Srivastava, D. R., Andrianov, V. M., and Piruzian, E. S. 1989. Tissue culture and plant regeneration of watermelon (Citrullus vulgaris Schrad, cv. Melitopolski.). Plant Cell Rep. 8: 300-302. Tang, W., Newton, R. J., and Weidner, D. A. 2007. Genetic transformation and gene silencing mediated by multiple copies of a transgene in eastern white pine. J. Exp. Bot. 58: 545-554. Tricoli, D. M., Carney, K. J., Russell, P. F., McMaster, J. R., Groff, D. W., Hadden, K. C., Himmel, P. T., Hubbard, J. P., Boeshore, M. L., and Quemada, H. D. 1995. Field evaluation of transgenic squash containing single or multiple virus coat protein gene constructs for resistance to cucumber mosaic virus, watermelon mosaic virus 2, and zucchini yellow mosaic virus. Bio/Technology 13: 1458-1465. Tsai, W. S., Shih, S. L., Green, S. K., and Jan, F. J. 2007. Occurrence and molecular characterization of Squash leaf curl Philippines virus in Taiwan. Plant Dis. 91: 907. Ugaki, M., Tomiyama, M., Kakutani, T., Hidaka, S., Kiguchi, T., Nagata, R., Sato, T., Motoyoshi, F., and Nishiguchi, M. 1991. The complete nucleotide sequence of cucumber green mottle mosaic virus (SH strain) genomic RNA. J. Gen. Virol. 72: 1487-1495. Wang, H., and Stubbs, G. 1994. The structure of cucumber green mottle mosaic virus at 3.4 Å resolution by x-ray fiber diffraction: significance for the evolution of tobamoviruses. J. Mol. Biol. 239: 371-384. Wang, S. M., and Chen, M. J. 1985. New strain of cucumber mottle mosaic virus causing mosaic symptoms on bottlegourd in Taiwan. Plant Prot. Bull. 27: 105-110. Yalcin-Mendia, N. Y., Ipek, M., Kacana, H., Curukb, S., Saria, N., Cetinerc, S., and Gabad, V. 2003. A histological analysis of regeneration in watermelon. J. Plant Biochem. Biotechnol. 12: 147-150. Yeh, S. D., and Chang, T. F. 1995. Nucleotide sequence of the N gene of watermelon silver mottle virus, a proposed new member of the genus Tospovirus. Phytopathology 85: 58-64. Yeh, S. D., Lin, Y. C., Cheng, Y. H., Jih, C. L., Chen, M. J., and Chen, C. C. 1992. Identification of Tomato Spotted Wilt-Like Virus on Watermelon in Taiwan. Plant Dis. 76: 835-840. Yeh, S. D., and Gonsalves, D. 1984. Purification and immunological analyses of cylindrical-inclusion protein induced by papaya ringspot virus and watermelon mosaic virus 1. Phytopathology 74: 1273-1278. Yoshioka, K., Hanada, K., Nakazaki, Y., Minobe, Y., Yakuwa, T., and Oosawa, K. 1992. Successful transfer of the cucumber mosaic virus coat protein gene to Cucumis melo L. Jpn. J. Breed. 42: 277-285. Yoshioka, K., Hanada, K., Harada, T., Minobe, Y., and Oosawa, K. 1993. Virus resistance in transgenic melon plants that express the cucumber mosaic virus coat protein gene and in their progeny. Jpn. J. Breed. 43: 629-634.
西瓜是台灣重要的經濟作物,但在栽培期間易受到多種植物病毒的侵害。在先前的研究中顯示當轉基因為silencer DNA連接多個病毒基因片段時可以藉由轉錄後基因沉寂機制提供對多種病毒的抗性,在本研究中使用相似的策略,藉由基因沉寂機制發展出可抗多種病毒的的轉基因西瓜,在本研究中將帶有西瓜銀斑病毒 (Watermelon silver mottle virus, WSMoV) 部份N基因做為silencer DNA其後連接胡瓜嵌紋病毒 (Cucumber mosaic virus, CMV)、胡瓜綠斑嵌紋病毒 (Cucumber green mottle mosaic virus, CGMMV) 及西瓜嵌紋病毒 (Watermelon mosaic virus, WMV) 部份鞘蛋白基因的表現卡匣構築於篩選標示基因與目標基因位於同一T-DNA或不同T-DNA的轉殖載體上,藉由農桿菌轉殖法進行西瓜基因轉殖,利用PCR檢測篩選標示基因,共獲得25個可增幅出篩選標示基因專一片段的基因株系,並利用南方雜合法確認目標基因嵌入西瓜基因組中不同的位置。所獲得的轉基因西瓜分別以CMV、CGMMV、WMV或混合三種病毒進行挑戰接種,評估其對於病毒的抗性,總共25個株系進行接種測試,其中有三個株系顯現出對CGMMV及WMV的抗性,這三個對CGMMV及WMV的抗病株系中有兩個對於CMV及混合接種也具有抗性。利用北方雜合法分析轉基因西瓜植株,可以觀察到抗病表現與低目標基因RNA轉錄累計是有相關的。我們的結果顯示當轉基因西瓜帶有不同的病毒鞘蛋白基因片段可以提供對多種病毒的抗性。目前已獲得由R0自交而得的R1種子,將評估R1植株對於CMV、CGMMV、WMV及WSMoV的廣泛抗性。

Watermelon is an important commercial crop in Taiwan and is prone to attack by several plant viruses. Previous studies have shown that transgenic plants with multiple viral gene fragments linked to an universal silencer DNA can provide resistance to multiple virus via posttranscriptional gene silencing (PTGS). In this study, similar approach was used to develop transgenic watermelon plants resistant to three different viruses via gene silencing. Transformation constructs with selection marker gene and targer gene either in the same T-DNA or in different T-DNAs containing a silencer DNA from the partial N gene of Watermelon silver mottle virus (WSMoV) fused to the partial coat protein genes of Cucumber mosaic virus (CMV), Cucumber green mottle mosaic virus (CGMMV) and Watermelon mosaic virus (WMV) were generated and used to transform watermelon plant via Agrobacterium-mediated transformation. A total of twenty-five trnasgenic lines were generated and confirmed by PCR amplification of selection marker gene. Southern blot was used for analyzing the copy number of the target gene inserted the genome of each transgenic watermelon plant. Transgenic watermelon plants were challenged with CMV, CGMMV, WMV or a mixture of these three viruses to evaluate the multiple virus resistance. Among the twenty-five tested lines, three lines showed resistant to CGMMV and WMV. Among these three resistant lines, two were also resistant to CMV and mixed inoculation of three viruses. The result of northern blotting of transgenic watermelon plants showed a correlation between the resistant phenotype and low accumulation level of target gene RNA transcripts. Our preliminary results indicate that transgenic watermelon carrying the fusion of different viral CP gene fragments can provide multiple virus resistance. R1 seeds from the self-pollinated resistant R0 plants were obtained and will be used to evaluate the broad-spectrum resistance to CMV, CGMMV, WMV and WSMoV.
其他識別: U0005-2208200700270800
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