低温条件下喷施乙烯利对温敏核不育水稻育性的影响
|
摘要
【目的】为解决我国南方温敏核不育水稻制种过程中遇低温易发生育性转换问题,对处于育性转换敏感期的温敏核不育水稻进行低温诱导并喷施不同浓度乙烯利,探讨其对温敏核不育水稻育性的影响。【方法】利用温敏核不育水稻株1S和准S为材料,在育性敏感时期对其进行连续7 d冷灌池低温(22.5℃)诱导并分别喷施乙烯合成促进物质乙烯利和抑制物质二氧化氯溶液,统计花粉可染率及套袋自交结实率,对乙烯合成关键酶进行实时荧光定量PCR分析。分离并分析了乙烯合成途径关键酶OsA CO5和Os ACS5的启动子。【结果】在22.5℃低温诱导条件下,株1S和准S连续7 d喷水处理后的可染花粉率分别是4.3%和8.9%,套袋结实率分别是1.6%和2.5%;与喷水对照相比,喷施1600 mg/L乙烯利,株1S和准S可染花粉率分别下降3.3和7.2个百分点,且都无结实;喷施1500 mg/L二氧化氯溶液的株1S和准S可染花粉率分别上升3.5和3.3个百分点,套袋结实率分别上升0.78和0.52个百分点。在低温诱导条件下(以喷水为对照),连续7 d喷施1600 mg/L乙烯利时,株1S和准S幼穗中OsA CO5和OsA CS5基因表达量显著升高,分别是对照(喷水)的1.19、1.24(株1S)和1.24、1.1(准S)倍;连续7 d喷施1500 mg/L二氧化氯溶液时,株1S和准S幼穗中Os ACO5基因表达量变化不显著,而株1S和准S中OsA CS5基因表达量显著降低,分别是对照(喷水)的93%和89%。与正常大田温度相比,在22.5℃低温诱导7 d时,喷水、喷施1600 mg/L乙烯利及1500 mg/L二氧化氯溶液处理分别会导致株1S和准S幼穗中OsA CO5和Os ACS5基因下调表达。此外,发现OsA CO5和OsA CS5的启动子除含TATA盒和CAAT盒等基本结构以外,还含有多个组织特异表达及乙烯调控顺式作用元件。【结论】低温诱导条件下,在温敏核不育水稻育性敏感期,喷施1600 mg/L乙烯利可抑制育性转换。
【Objective】This research aims to elucidate the mechanism behind sterility conversion(infertile convert into fertile) in low temperature of thermo-sensitive genic male sterile rice in seed production in South China by applying exogenous ethephon.【Method】The thermo-sensitive genic male sterile rice cultivars Zhu 1 S and Zhun S were treated with artificial cooling water tank(low temperature, 22.5℃) induction for 7 days and sprayed with ethephon(which promote synthesis of ethylene) and the chlorine dioxide solution(inhibitory substances) during fertility-sensitive period. Then the pollen dyeing rate and the bagged seed-setting rate were calculated and the quantitative real-time PCR was conducted. In addition, the promoters of OsA CO5 and OsA CS5, which are key enzymes of ethylene synthesis pathway, were isolated and analyzed.【Result】Under 22.5℃ for 7 days, the pollen rate of Zhu 1 S and Zhun S was 4.3% and 8.9%, and bagged seed-setting rate was 1.6% and 2.5%, respectively. Compared with the water spray control, the pollen rate of lines Zhu 1 S and Zhun S was reduced by 3.3 and 7.2 percent, respectively when spraying with 1600 mg/L ethephon. When spraying with 1500 mg/L of chlorine dioxide solution, the pollen rate was increased by 3.5 and 3.3 percent, and the bagged seed setting rate increased by 0.78 and 0.52 percent, respectively. The expression of OsA CO5 and OsA CS5 in young panicles was significantly increased at 1600 mg/L of ethephon under 22.5℃ induction for 7 days, which were 1.19, 1.24(Zhu 1 S) and 1.24, 1.1(Zhun S) times of that of the control(water spray) respectively. When sprayed with 1500 mg/L of chlorine dioxide solution for 7 days, the expression of OsA CS5 gene in the Zhu 1 S and Zhun S was significantly reduced, which were 0.93 and 0.89 times of that of the control(water spray), respectively, but the expression of OsA CO5 was not significantly changed. Compared with normal field temperature(control) for 7 days, water spraying, 1600 mg/L of ethephon and 1500 mg/L of chlorine dioxide solution at 22.5℃ down-regulated the expression of Os ACO5 and OsA CS5 in young panicles of Zhu 1 S and Zhun S. The promoters of OsA CO5 and OsA CS5 contained multiple tissue-specific expression elements and ethylene-regulated cis-acting elements in addition to the basic structures such as TATA-box and CAAT-box. 【Conclusion】Under low temperature condition, spraying 1600 mg/L ethephon in the sensitive period of thermo-sensitive genic male sterile rice had inhibitory effect on fertility conversion from sterility to fertility.
【Objective】This research aims to elucidate the mechanism behind sterility conversion(infertile convert into fertile) in low temperature of thermo-sensitive genic male sterile rice in seed production in South China by applying exogenous ethephon.【Method】The thermo-sensitive genic male sterile rice cultivars Zhu 1 S and Zhun S were treated with artificial cooling water tank(low temperature, 22.5℃) induction for 7 days and sprayed with ethephon(which promote synthesis of ethylene) and the chlorine dioxide solution(inhibitory substances) during fertility-sensitive period. Then the pollen dyeing rate and the bagged seed-setting rate were calculated and the quantitative real-time PCR was conducted. In addition, the promoters of OsA CO5 and OsA CS5, which are key enzymes of ethylene synthesis pathway, were isolated and analyzed.【Result】Under 22.5℃ for 7 days, the pollen rate of Zhu 1 S and Zhun S was 4.3% and 8.9%, and bagged seed-setting rate was 1.6% and 2.5%, respectively. Compared with the water spray control, the pollen rate of lines Zhu 1 S and Zhun S was reduced by 3.3 and 7.2 percent, respectively when spraying with 1600 mg/L ethephon. When spraying with 1500 mg/L of chlorine dioxide solution, the pollen rate was increased by 3.5 and 3.3 percent, and the bagged seed setting rate increased by 0.78 and 0.52 percent, respectively. The expression of OsA CO5 and OsA CS5 in young panicles was significantly increased at 1600 mg/L of ethephon under 22.5℃ induction for 7 days, which were 1.19, 1.24(Zhu 1 S) and 1.24, 1.1(Zhun S) times of that of the control(water spray) respectively. When sprayed with 1500 mg/L of chlorine dioxide solution for 7 days, the expression of OsA CS5 gene in the Zhu 1 S and Zhun S was significantly reduced, which were 0.93 and 0.89 times of that of the control(water spray), respectively, but the expression of OsA CO5 was not significantly changed. Compared with normal field temperature(control) for 7 days, water spraying, 1600 mg/L of ethephon and 1500 mg/L of chlorine dioxide solution at 22.5℃ down-regulated the expression of Os ACO5 and OsA CS5 in young panicles of Zhu 1 S and Zhun S. The promoters of OsA CO5 and OsA CS5 contained multiple tissue-specific expression elements and ethylene-regulated cis-acting elements in addition to the basic structures such as TATA-box and CAAT-box. 【Conclusion】Under low temperature condition, spraying 1600 mg/L ethephon in the sensitive period of thermo-sensitive genic male sterile rice had inhibitory effect on fertility conversion from sterility to fertility.
引文
[1]袁隆平.发展杂交水稻保障粮食安全.杂交水稻,2010,25(S1):1-2.Yuan L P.Development of hybrid rice to ensure food security.Hybrid Rice,2010,25(S1):1-2.(in Chinese with English abstract)
[2]乔娟.中国主要粮食国际竞争力研究.北京:中国农业科学院,2004.Qiao J.The study on international competitiveness of China's main grain.Beijing:Chinese Academy of Agricultural Sciences,2004.(in Chinese with English abstract)
[3]陈正洪,袁业畅.武汉盛夏低温及其对农业的影响初析.湖北农业科学,1990,36,4(8):9-12.Chen Z H,Yuan Y C.The preliminary analysis of low temperature in summer in Wuhan and its influence on Agriculture.Hubei Agric Sci,1990,36,4(8):9-12.(in Chinese)
[4]陈大洲,王任初,熊焕金,陈春莲.几个籼型光敏感核不育系生产应用简介.江西农业科技,1991(3):12-13.Chen D Z,Wang R C,Xiong H J,Chen C L.A brief introduction to the production and application for several indica photoperiod sensitive genic male sterile lines.Jiangxi Agric Sci Technol,1991(3):12-13.(in Chinese with English abstract)
[5]Salgotral R K,Gupta B B,Ahmed M I.Characterization of thermo-sensitive genic male sterility(TGMS)rice genotypes(Oryza sativa L.)at different altitudes.Aus JCrop Sci,2012;6(6):957-962.
[6]周广洽,李训贞,谭周鎡.水稻籼型不育系安农S-1不育基因表达条件和育性转换规律的研究.湖南师范大学:自然科学学报,1990,13(4):365-372.Zhou G Q,Li X Z,Tan Z Z.Study on the expression conditions of indica male sterile lines rice gene S-1 and fertility transformation.J Hunan Normal Univ:Nat Sci,1990,13(4):365-372.(in Chinese with English abstract)
[7]陈良碧,周广洽.温光条件对水稻安农s-I/衡农S-l的育性及生理的影响.植物学报,1994,36(S):119-1234Chen L B,Zhou G Q.The effect on sterility and physiology of Annong s-I/Hengnong S-l under temperature and light characteristics.J Plant Bot,1994,36(S):119-123.(in Chinese with English abstract)
[8]刘忠奇.光温敏核不育水稻育性转换生理及相关蛋白的研究.长沙:湖南农业大学,2013.Liu Z Q.Studying on physiology and related proteins of the fertility transformation in thermo-sensitive genic male sterile rice.Changsha:Hunan Agricultural University,2013.(in Chinese with English abstract)
[9]Song L R,Liu Z Q,Tong J H,Xiao L T,Ma H,Zhang HQ.Comparative proteomics analysis reveals the mechanism of fertility alternation of thermosensitive genic male sterile rice lines under low temperature inducement.Proteomics,2015(15):1884-1905.
[10]Zhou Y F,Zhang X Y,Xue Q Z.Fine mapping and candidate gene prediction of the photoperiod and thermo-sensitive genic male sterile gene pms1(t)in rice.Biomed&Biotechnol,2011,12(6):436-447.
[11]Lival K J,Schmittgen T D.Analysis of relative gene expression data using real-time quantitative PCR and the2-??CT method.Methods,2001,25:402-408.
[12]袁隆平,陈洪新.杂交水稻育种栽培学.长沙:湖南科学技术出版社,1988:364Yuan L P.Chen H X.Hybrid Rice Breeding and Cultivation.Changsha:Hunan Science and Technology Press,1988:364.(in Chinese)
[13]斯华敏,付亚萍,刘文真,孙宗修,胡国成.水稻光温敏雄性核不育系的系谱分析.作物学报,2012,38(3):394-407.Si H M,Fu Y P,Liu W Z,Sun Z X,Hu G C.Pedigree analysis of photoperiod-thermo sensitive genic male sterile rice.Acta Agron Sin,2012,38(3):394-407.(in Chinese with English abstract)
[14]邓华凤,舒福北,袁定阳.安农S-1的研究及其利用概况.杂交水稻.1999,14(3):1-3Deng F H,Shu F B,Yuan D Y.Research and utilization of Annong S-1.Hybrid Rice,1999,14(3):1-3.(in Chinese with English abstract)
[15]段敏,谢留杰,黄善军,潘晓飚.水稻籼型不育系选育研究进展.中国稻米,2016,22(3):25-29.Duan M,Xie L J,Huang S J,Pan X B.Advances in breeding of indica CMS lines rice.China Rice,2016,22(3):25-29.(in Chinese with English abstract)
[16]Johnson P R,Ecker J R.The ethylene gas signal transduction pathway:A molecular perspective.Ann Rev Genet,1998,32,227-254.
[17]Abeles F B,Morgan P W,Saltveit M E.Ethylene in Plant Biology.New York:Academic Press,1992:1-296.
[18]王熹,阐瑞芬,蒋志荣,徐马福.乙烯利诱导水稻雄性不育的生理效应.植物生理学报,1981,7(1):11-18.Wang X,Chan R F,Jiang Z R,Xu M F.Ethrel induced male sterile rice physiological effect.J Plant Physiol,1981,7(1):11-18.(in Chinese with English abstract)
[19]李凝.乙烯的生物合成及其在农业上的应用//中国科学技术协会.生命科学与生物技术中国科协第三届青年学术年会论文集.北京:中国科学技术出版社,1998.Li N.Ethylene biosynthesis and its application in agriculture//China Association for Science and Technology.Life Science and Biotechnology:The Third Annual Academic Proceedings of China Association for Science and Technology.Beijing:China Science and Technology Press,1998.(in Chinese with English abstract)
[20]陈知子.植物化学杀雄研究进展.资源开发与市场,2010(2):143-145.Chen Z Z.Advances in the study of plant chemical killing.Res Dev&Market,2010(2):143-145.(in Chinese)
[21]刘娜.小麦光温敏雄性不育系孤雌生殖和育性调节研究.北京:首都师范大学,2007.Liu N.The studies on parthenogenesis and fertility regulation of photoperiod-thermo-sensitive genic male sterile lines wheat.Beijing:Capital Normal University,2007.(in Chinese with English abstract)
[22]谢学民,张全德,朱汉如.小麦化学杀雄制种技术.农业科技通讯,1979(5):5-6Xie X M,Zhang Q D,Zhu H R,Chemical male sterile seed production technique in Wheat.Agric Sci&Technol Newsl,1979(5):5-6.(in Chinese)
[23]肖层林,张海清,麻浩.植物杂种优势原理与利用.北京:高等教育出版社,2015:52-53.Xiao C L,Zhang H Q,Ma H.Principle and utilization of plant heterosis.Beijing:Higher Education Press,2015:52-53.(in Chinese)
[24]谢学民,张全德,朱汉如,刘愚.14C-乙烯利对小麦杀雄机理的研究:14C-乙烯和14C-乙烯利的吸收、分布和代谢.浙江农业大学学报,1980(2):33-41.Xie Q M,Zhang Q D,Zhu H R,Liu Y.Study on the mechanism of 14C-ethephon on wheat chemical hybridizing:Absorption,distribution and metabolism of14C-ethylene and 14C-ethephon.J Zhejiang Agric Univ,1980(2):33-41.(in Chinese with English abstract)
[25]骆炳山,李文斌.湖北光敏核不育水稻育性转换机理初探.华中农业大学学报,1990,9(1):7-12Luo B S,Li W B.Preliminary study on the mechanism of fertility conversion of photoperiod sensitive genic male sterile rice in Hubei.J Huazhong Agric Univ,1990,9(1):7-12.(in Chinese with English abstract)
[26]Lau O L,Yang S F.Inhibition of ethylene production by cobaltousin.Plant Physiol,1976,58(1):114-117.
[27]李德红,骆炳山,屈映兰.光敏核不育水稻幼穗的乙稀生成与育性转换.植物生理学报,1996,22(3):320-326.Li D H,Luo B S,Qu Y L.Ethylene generation and fertility transformation in young panicle of photoperiod sensitive genic male sterile rice.J Plant Physiol,1996,22(3):320-326.(in Chinese with English abstract)
[2]乔娟.中国主要粮食国际竞争力研究.北京:中国农业科学院,2004.Qiao J.The study on international competitiveness of China's main grain.Beijing:Chinese Academy of Agricultural Sciences,2004.(in Chinese with English abstract)
[3]陈正洪,袁业畅.武汉盛夏低温及其对农业的影响初析.湖北农业科学,1990,36,4(8):9-12.Chen Z H,Yuan Y C.The preliminary analysis of low temperature in summer in Wuhan and its influence on Agriculture.Hubei Agric Sci,1990,36,4(8):9-12.(in Chinese)
[4]陈大洲,王任初,熊焕金,陈春莲.几个籼型光敏感核不育系生产应用简介.江西农业科技,1991(3):12-13.Chen D Z,Wang R C,Xiong H J,Chen C L.A brief introduction to the production and application for several indica photoperiod sensitive genic male sterile lines.Jiangxi Agric Sci Technol,1991(3):12-13.(in Chinese with English abstract)
[5]Salgotral R K,Gupta B B,Ahmed M I.Characterization of thermo-sensitive genic male sterility(TGMS)rice genotypes(Oryza sativa L.)at different altitudes.Aus JCrop Sci,2012;6(6):957-962.
[6]周广洽,李训贞,谭周鎡.水稻籼型不育系安农S-1不育基因表达条件和育性转换规律的研究.湖南师范大学:自然科学学报,1990,13(4):365-372.Zhou G Q,Li X Z,Tan Z Z.Study on the expression conditions of indica male sterile lines rice gene S-1 and fertility transformation.J Hunan Normal Univ:Nat Sci,1990,13(4):365-372.(in Chinese with English abstract)
[7]陈良碧,周广洽.温光条件对水稻安农s-I/衡农S-l的育性及生理的影响.植物学报,1994,36(S):119-1234Chen L B,Zhou G Q.The effect on sterility and physiology of Annong s-I/Hengnong S-l under temperature and light characteristics.J Plant Bot,1994,36(S):119-123.(in Chinese with English abstract)
[8]刘忠奇.光温敏核不育水稻育性转换生理及相关蛋白的研究.长沙:湖南农业大学,2013.Liu Z Q.Studying on physiology and related proteins of the fertility transformation in thermo-sensitive genic male sterile rice.Changsha:Hunan Agricultural University,2013.(in Chinese with English abstract)
[9]Song L R,Liu Z Q,Tong J H,Xiao L T,Ma H,Zhang HQ.Comparative proteomics analysis reveals the mechanism of fertility alternation of thermosensitive genic male sterile rice lines under low temperature inducement.Proteomics,2015(15):1884-1905.
[10]Zhou Y F,Zhang X Y,Xue Q Z.Fine mapping and candidate gene prediction of the photoperiod and thermo-sensitive genic male sterile gene pms1(t)in rice.Biomed&Biotechnol,2011,12(6):436-447.
[11]Lival K J,Schmittgen T D.Analysis of relative gene expression data using real-time quantitative PCR and the2-??CT method.Methods,2001,25:402-408.
[12]袁隆平,陈洪新.杂交水稻育种栽培学.长沙:湖南科学技术出版社,1988:364Yuan L P.Chen H X.Hybrid Rice Breeding and Cultivation.Changsha:Hunan Science and Technology Press,1988:364.(in Chinese)
[13]斯华敏,付亚萍,刘文真,孙宗修,胡国成.水稻光温敏雄性核不育系的系谱分析.作物学报,2012,38(3):394-407.Si H M,Fu Y P,Liu W Z,Sun Z X,Hu G C.Pedigree analysis of photoperiod-thermo sensitive genic male sterile rice.Acta Agron Sin,2012,38(3):394-407.(in Chinese with English abstract)
[14]邓华凤,舒福北,袁定阳.安农S-1的研究及其利用概况.杂交水稻.1999,14(3):1-3Deng F H,Shu F B,Yuan D Y.Research and utilization of Annong S-1.Hybrid Rice,1999,14(3):1-3.(in Chinese with English abstract)
[15]段敏,谢留杰,黄善军,潘晓飚.水稻籼型不育系选育研究进展.中国稻米,2016,22(3):25-29.Duan M,Xie L J,Huang S J,Pan X B.Advances in breeding of indica CMS lines rice.China Rice,2016,22(3):25-29.(in Chinese with English abstract)
[16]Johnson P R,Ecker J R.The ethylene gas signal transduction pathway:A molecular perspective.Ann Rev Genet,1998,32,227-254.
[17]Abeles F B,Morgan P W,Saltveit M E.Ethylene in Plant Biology.New York:Academic Press,1992:1-296.
[18]王熹,阐瑞芬,蒋志荣,徐马福.乙烯利诱导水稻雄性不育的生理效应.植物生理学报,1981,7(1):11-18.Wang X,Chan R F,Jiang Z R,Xu M F.Ethrel induced male sterile rice physiological effect.J Plant Physiol,1981,7(1):11-18.(in Chinese with English abstract)
[19]李凝.乙烯的生物合成及其在农业上的应用//中国科学技术协会.生命科学与生物技术中国科协第三届青年学术年会论文集.北京:中国科学技术出版社,1998.Li N.Ethylene biosynthesis and its application in agriculture//China Association for Science and Technology.Life Science and Biotechnology:The Third Annual Academic Proceedings of China Association for Science and Technology.Beijing:China Science and Technology Press,1998.(in Chinese with English abstract)
[20]陈知子.植物化学杀雄研究进展.资源开发与市场,2010(2):143-145.Chen Z Z.Advances in the study of plant chemical killing.Res Dev&Market,2010(2):143-145.(in Chinese)
[21]刘娜.小麦光温敏雄性不育系孤雌生殖和育性调节研究.北京:首都师范大学,2007.Liu N.The studies on parthenogenesis and fertility regulation of photoperiod-thermo-sensitive genic male sterile lines wheat.Beijing:Capital Normal University,2007.(in Chinese with English abstract)
[22]谢学民,张全德,朱汉如.小麦化学杀雄制种技术.农业科技通讯,1979(5):5-6Xie X M,Zhang Q D,Zhu H R,Chemical male sterile seed production technique in Wheat.Agric Sci&Technol Newsl,1979(5):5-6.(in Chinese)
[23]肖层林,张海清,麻浩.植物杂种优势原理与利用.北京:高等教育出版社,2015:52-53.Xiao C L,Zhang H Q,Ma H.Principle and utilization of plant heterosis.Beijing:Higher Education Press,2015:52-53.(in Chinese)
[24]谢学民,张全德,朱汉如,刘愚.14C-乙烯利对小麦杀雄机理的研究:14C-乙烯和14C-乙烯利的吸收、分布和代谢.浙江农业大学学报,1980(2):33-41.Xie Q M,Zhang Q D,Zhu H R,Liu Y.Study on the mechanism of 14C-ethephon on wheat chemical hybridizing:Absorption,distribution and metabolism of14C-ethylene and 14C-ethephon.J Zhejiang Agric Univ,1980(2):33-41.(in Chinese with English abstract)
[25]骆炳山,李文斌.湖北光敏核不育水稻育性转换机理初探.华中农业大学学报,1990,9(1):7-12Luo B S,Li W B.Preliminary study on the mechanism of fertility conversion of photoperiod sensitive genic male sterile rice in Hubei.J Huazhong Agric Univ,1990,9(1):7-12.(in Chinese with English abstract)
[26]Lau O L,Yang S F.Inhibition of ethylene production by cobaltousin.Plant Physiol,1976,58(1):114-117.
[27]李德红,骆炳山,屈映兰.光敏核不育水稻幼穗的乙稀生成与育性转换.植物生理学报,1996,22(3):320-326.Li D H,Luo B S,Qu Y L.Ethylene generation and fertility transformation in young panicle of photoperiod sensitive genic male sterile rice.J Plant Physiol,1996,22(3):320-326.(in Chinese with English abstract)