臺灣博碩士論文加值系統

摘要
水稻（Oryza sativa L.）是目前重要的單子葉模式植物，對於水稻特殊性狀的選拔以及基因定位等研究，利用近同源系（near-isogenic line）是相當快速的方法；本試驗即是利用187個微衛星標誌（microsatellite marker），在Shiokari與其矮性近同源系d6為親本的F2族群中，尋找與d6矮性基因緊密連鎖的標誌，結果在水稻第七條染色體的上端，找到四個有簡單序列長度多型性（simple sequence length polymorphism）的標誌，此與目標性狀有連鎖的標誌，可在標誌輔助選拔 ( marker-assisted selection ) 上應用；未來將進一步進行連鎖分析以及據圖選殖的研究。

關鍵字
水稻、近同源系、微衛星標誌、簡單序列重複、簡單序列長度多型性、Shiokari、標誌輔助選拔

Abstract

Rice is an important monocotyledones model plant, near-isogenic lines were very useful tool to selection rice special trait or genome. In this study, to tag the precise location of d6 locus, 187 microsatellite markers were mapped on the F2 population derived from the cross between Shiokari and the near iosgenic line d6.
The result indicated that four SSR markers on the top of rice chromosome 7 were linked to d6 locus, which can facilitate the application of marker-assisted selection and map-base cloning in the future.
Key words: rice, ( Oryza sativa L.) , near isogenic lines ( NILs ), microsatellite marker, simple sequence repeat (SSR), simple sequence length polymorphism ( SSLP ), Shiokari, marker-assisted selection ( MAS ).

目錄
頁次
中文摘要………………………………………………………………..Ⅴ
英文摘要………………………………………………………………..Ⅵ
壹、前人研究……………………………………………………………..1
ㄧ、水稻基因組之研究……………………………………………..1
二、植物矮型表現型之研究………………………………………..1
三、簡單重複序列標誌之研究…………………………………….3
四、近同源系之研究……………………………………………….4
五、截切放大多型性序列標誌之研究……………………………..5
六、功能性標誌之研究……………………………………………6
貳、材料與方法…………………………………………………………7
一、試驗材料………………………………………………………7
二、植株外表型性狀調查…………………………………………7
三、水稻葉片DNA萃取…………………………………………..7
四、水稻SSR分子標誌分析………………………………………8
五、CAPS標誌的設計……………………………………………..8
六、聚丙醯胺膠體電泳和銀染法………………………………….9
七、FM的設計……………………………………………………10
八、Gelelution……………………………………………………10
九、F2子代株高統計分析…………………………………………11
十、F2性狀相關分析………………………………………………11
參、結果…………………………………………………………………12
一、植株外表型性狀調查…………………………………………12
二、F2子代株高數量分群…………………………………………12
三、SSR分子標誌篩選矮生近同源系……………………………12
四、SSR分子標誌篩選親本………………………………………12
五、F2族群利用有多型性的SSR標誌分析………………………13
六、CAPS標誌的分析…………………………………………….13
七、FM的分析……………………………………………………..13
肆、討論…………………………………………………………………14
伍、結語…………………………………………………………………19
陸、參考文獻……………………………………………………………20

吳詩都與曾富生。1984, 水稻矮性基因對農藝性狀之作用。中華農學會報新第126期:10-18。
李長沛。1998, 栽培稻（O.sativa）與野生稻（O.nivara）種間雜交種後代的遺傳及育種行為。國立中興大學農藝學研究所碩士論文。
Andersen, J. R., and Thomas, L. 2003. Functional markers in plants. Trends in plant Sci. 8:554-560.
Andrzef, K., and Ausubel, F. M. 1993. A procedure for mapping Arabidopsis mutations using co-dominant ecotype-specific PCR-based markers. The Plant J. 4:403-410.
Ashikari, A., Jianzhong, W., Masahiro, Y., Yakuji, S., and Atsushi, Y. 1999. Rice gibberellin-insensitive dwarf mutant gene Dwarf1 encodes the α-subunit of GTP-binding protein. Proc. Natl. Acad. Sci. USA 96:10284-10289.
Ashikari, M., Itoh, H., Ueguchi-Tanaka, M., Akie, S., Kenji, G., Hidemi, K., and Makoto, M. 2003. Gibberellin signal transduction in rice. J. Plant Growth Trgul. 22:141-151.
Ayre, N. M., NcClung, A. M., Larkin, P. D., Bligh, H. F. J., Jones, C. A., and Park, W. D. 1997. Microsatellites and a single-nucleotide polymorphis differentiate apparent amylase classes in an extended pedigree of US rice germ plasm. Theor.Appl. Genet. 94:773-781.
Bassam, B. J., Caetano-Anolles, G., and Gresshoff, P. M. 1991. Fast and sensitive silver staining of DNA in polyacrylamide gels. Anal. Biochem. Jul.196: 80-83.
Bligh, H. F. J., Blackhall, N. W., Edwards, K. J., and McClung, A. M. 1999. Using amplified fragment length polymorphisms and simple sequence length polymorphisms to identify cultivars of brown and white milled rice. Crop Sci. 39:1715-1721.
Blair, M. W., and McCouch, S. R. 1997. Microsatellite and sequence-taged site markers diagnostic for the rice bacterial leaf blight resistance gene xa-5. Theor. Appl. Genet. 95:174-184.
Chen, X., Temnykh, S., Xu, Y., Cho, Y. G., and McCouch, S.R. 1997. Development of a microsatellite framework map providing genome-wide coverage in rice ( Oryza sativa L. ) Theor. Appl. Genet. 95:553-567
Chono, M., Ichiro, H., Haruko, Z., Koichi, Y., Daisuke, S., Kazuyoshi, T., Suguru, T., Tsuguhiro, H., and Yoshiaki, W. 2003. A semidwarf phenotype of barley uzu results from a nucleotide substitution in the gene encoding a putative brassinosteroid receptor. Plant Physiol. 133:1209-1219.
Cho, T.G., Ishii, T., Temnykh, S., Chen, X., Lipovich, L., McCouch, S. R., Park, W. D., Ayres, N., and Cartinhour, S. 2000. Diversity of microsatellites derived from genomic libraries and GenBank sequences in rice ( Oryza sativa L. ) Theor. Appl. Genet. 100:713-722.
Cho, Y. G., Eun, M. Y., NcCouch, S.R., and Chae, Y. A. 1994. The semidwarf gene, sd-1, of rice(Oryza sativa L.) Ⅱ. Molecular mapping and marker-assisted selection. Thior. Appl. Genet. 89:54-59.
Eujayl, I., Sorrells, M. E., Baum, M., Wolters, P., and Powell, W. 2002. Isolation of EST-derived microsatellite markers for genotyping the A and B genomes of wheat. Theor. Appl. Genet. 104:399-407.
Fridborg, I., Sandra, K., Thomas, M., and Eva, S. 1999. The Arabidopsis dwarf mutant shi exhibits reduced gibberellin responses conferred by overexpression of a new putative zinc finger trotein. Plant Cell 11:1019-32.
Harberd, N. P., and Michael, F. 1989.Genetics of dominant gibberelllin-insensitive dwarfism in maize. Genetics 121:827-838.
Hayden, M. J., and Sharp, P. J. 2001 Targeted development of informative microsatellite (SSR) markers. Nucle. Acids Res. 29:8e44.
Itoh, H., Ueguchi-Tanaka, M., Sakamoto, T., Kayano, T., Tanaka, H., Ashikari, M., and Matsuoka, M. 2002. Modification of rice plant height by suppressing the height-controlling gene, D18, in rice. Breed. Sci. 52:215-218.
Ishimaru, K., Ono, K., and Kashiwagi, T. 2004. Identification of a new gene controlling plant height in rice using the candidate-gene strategy. Planta 218:388-395.
Juliano, B. O. 1972. Physico-chemical properties of starch and protein in relation to grain quality and nutritional value of rive.
Junjian, N., Peter, M C., and David J. M. 2002. Evaluation of genetic diversity in rice subspecies using microsatellite markers. Crop Sci. 42:601-607.
Keller, B., and Feuillet, C. 2000. Colinearity and gene density in grass genomes. Trends Plant Sci. 5:246-251.
Kobayashi, M., Paul, G., Clive, R. S., Bernard, O. P., and Jake, M. 1994.The metabolism of gibberellin A20 to gibberellin A1 by tall and dwarf mutants of Oryza sative and Asabidopsis thaliana. Plant Physiol. 106:1367-1372.
Konovalov, F., Eugenia, T., and Sergei, G. 2005. A CAPS marker set for mapping in linkage groupⅢ of pea（Pisum sativum L.） Cellular&Molecular Biol. Lett. 10:163-171.
Liu, B., and Jonathan, F. W. 2002. Non-Mendelian phenomena in allopolyploid genome evolution. Curr. Genomics. 3:1-17
Martin, G. B., Williams, J. G. K., and Tanksley, S. D. 1991. Rapid identification of markers linked to a Pseudomonas resistance gene in tomato by using random primers and near-isogenic lines. Proc. Natl. Acad. Sci. USA. 88:2236-2340.
McCouch, S. R., Teytelman, L., Xu, Y., Lobos, K. B., Clare, K., Walton, M., Fu, B., Maghirang, R., Li, Z., Xing, Y., Zhang, Q., Kono, I., Yano, M., Fjellstrom, R., DeClerck, G., Schneider, D., Cartinhour, S., Ware, D., and Stein, L. 2002. Development and mapping of 2240 new SSR markers for rice ( Oryza sativa L. ). DNA Res. 9:199-207.
Milach, S. C. K., Phillips, R. L., Stuthman, D. D., and Monkawa, T. 1998.Inheritance of a new dwarfing gene in Oat. Crop Sci. 38:356-360.
Milach, S.C.K., Rines, H. W., and Phillips R. L. 2002. Plant Height components and gibberellic acid response of oat dwarf lines. Crop Sci. 42:1147-1154.
Mohan, M., Nair, S., Bhagwat, A., Krishna, T. G., Yano, M., Bhatia, C. R., and Sasaki, T. 1997. Genome mapping, molecular markers and marker-assisted selection in crop plants. Mol. Breed. 3:87-103.
Murai, M., N. Shinbashi, S. Sato, K. Sato, H. Araki and M. Ehara. 1995. Efect of the dwarfing gene from Dee-geo-woo-gen on culm and internode lengths, and its response to fertilizer in rice. Breed. Sci. 45:7-14.
Nelson, J. C. 1997. QGENE: software for marker-based genomic analysis and breeding. Mol. Breed. 3:239-245.
Panaud, O., Chen, X., and McCouch, S. R. 1996. Development of microsatellite markers and characterization of simple sequence length polymorphism (SSLP) in rice (Oryza sativa L.) Nol. Gen. Genet. 252:597-607.
Sasaki, A., Ashikari, M., Ueguchi-Tanaka, M., Itoh, H., Nishimura, A., Swapan, D., Ishiyama, K., Kitano, H., and Matsuoka, M. 2002. A mutant gibberellin-synthesis gene in rice. Nature 416:701.
Sasaki, T. and Benjamin B. 2000. International rice genome sequencing project: the effort to completely sequence the rice genome. Cur. Opin. Plant Biol. 3:138-141.
Saito, A., Yano, M. K., Nakagahra, M., Yoshimura, A., Saito, K., Kuhara, S., Ukai, Y., Kawase, M., Nagamine, T., Yoshimura, S., Ideta, O., Ohsawa, R., Hayano, Y., Iwata, N., and Sugiura, M. 1991. Linkage map of restriction fragment length polymorphism loci in rice. Jpn. P. Breed. 41:665-670.
SAS Institute ( 1999 ) SAS/STAT® User’s Guide, Version 8, SAS Institute, Cary, NC, USA.
Sarfatti, M., J. Katan, R. Fluhr and D. Zamir. 1989. An RFLP marker in tomato linked to the Fusarium oxysporum resistance gene I2. Theor. Appl. Genet. 78: 755-759.
Singh, S., Sidhu, J. S., Hunag, N., Vikal, Y., Li , Z., Brar, D. S., Dhaliwal, H. S., and Khush G. S. 2001.Pyramiding three bacterial blight resistance genes ( xa5, xa13 and Xa21 ) using marker-assisted selection into indica rice cultivar PR106. Theor. Appl. Genet. 102:1011-1015.
Spielmeyer, W., Marc, H. E., and Peter M. C. 2002. Semidwarf (sd-1), “green revolution” rice, contains a defective gibberellin 20-oxidase gene. Proc. Natl. Acad. Sci. USA 99:9043-9048.
Suh, H. S. and M.H. Heu. 1978. The segregation mode of plant height in the cross of rice varieties. Ⅱ. Linkage analysis of the semi-dwarfness of rice varienty “Tongil”. Korean J. Breed. 10:1-6.
Temnykh, S., Park, W. D., Ayres, N., Cartinhour, S., Hauck, N., Lipovich, L., Cho, Y. G., Ishii, T., and McCouch, S. R. 2000. Mapping and genome organization of microsatellite sequences in rice (Owyza sativa L.)Theor. Appl. Genet. 100:697-712.
Thiel, T., Michalek, W., Varshney, R. K., and Graner, A. 2003. Expliting EST atabases for the development and characterization of gene-derived SSR-markers in barley( Hordeum vulgare L.) Theor. Appl. Genet. 106:411-422.
Ware, D. and Stein, L. 2003. Comparison of genes among cereals. Curr. Opin. Plant Biol. 6:121-127
Weiland, J. J. and Yu, M. H. 2003. A cleaved amplified polymorphic sequence（CAPS） marker associated with root-knot nematode resistance in sugarbeet. Crop Sci 43:1814-1818.
Young, N. D., D. Zamir, M. W. Ganal and S. D. Tanksley. 1988. Use of isogenic lines and simultaneous probing to identify dna markers tightly linked to the Tm-2a gene in tomato. Genetics Society of America 120:579-585.
Zhang, Y., and Stommel, J. R. 2001. Development of SCAR and CAPS markers linked to the Beta gene in tomato. Crop Sci. 41:1602-1608.