γ射线诱变籼稻保持系T98B突变体的鉴定与农艺性状分析
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摘要
辐射诱变是农作物种质创新的重要手段。本研究以杂交稻骨干亲本T98B为研究对象,以300 Gy剂量的60Co-γ射线对其成熟种子进行处理,从辐射后代中鉴定出农艺性状变异材料,划分了变异类群,分析了各类型突变频率,为遗传改良奠定了材料基础。结果显示,经γ射线处理后的M1和M2种子的发芽率受到了显著抑制,分别比对照T98B降低了22. 35%和14. 67%;从M2群体中初步筛选出了207个变异单株;经M3～M5的遗传稳定性测试后最终获得了168份可稳定遗传的突变体,总突变频率达到了0. 673%。按照变异部位将突变体划分成叶变异、茎/蘖变异和穗粒/花变异等3种类群,各类群分别含有34份、37份和97份突变体,占比20. 24%、20. 02%和57. 74%;育性、叶色和株高等性状的突变频率较高,分别达到了0. 212%,0. 092%和0. 088%。此外,本文简要描述了各类群的表型变异特点。结果表明,利用γ射线诱变能产生丰富的农艺性状突变体,本研究为水稻遗传改良提供新的种质资源奠定了基础。
Radiation mutagenesis is available for crops germplasm creation. In this study,gamma-ray irradiation was used to induce the rice cultivar T98B which is a main maintainer line of hybrid rice,under the treatment of mature seeds at a dose of 300 Gy. It focused on screening and grouping of agronomic mutants,and the frequency for each type was also investigated. The results showed that,by contrast of T98B,the germination rate for M1 plants and M2 plants were both significantly suppressed,with a decrease of 22. 35% and 14. 67% respectively. In M2 population,207 candidate mutants were screened. Of which,168 were validated steadily inheritable via a phenotyping test from M3 to M5,accounting for a total mutation frequency of 0. 673%. Furthermore,they were divided into 3 groups regarding leaf group,stem/tiller group and grain/flower group,according to the part that it occurred to mutate on. Each group above included 34,37 and 97 mutants respectively,representing a ratio of 20. 24%,20. 02% and 57. 74%. The common mutants were the ones varied on fertility,leaf color and plant height,and it presented a mutation frequency of 0. 212%,0. 092%and 0. 088% respectively. Moreover,a brief description on the phenotypes for each group was provided. The results indicated that,agronomic mutants in rice could be effectively obtained via gamma-ray irradiation,and several mutants gained here would be severed as useful germ resources for advancing the work on rice genetic improvement.
Radiation mutagenesis is available for crops germplasm creation. In this study,gamma-ray irradiation was used to induce the rice cultivar T98B which is a main maintainer line of hybrid rice,under the treatment of mature seeds at a dose of 300 Gy. It focused on screening and grouping of agronomic mutants,and the frequency for each type was also investigated. The results showed that,by contrast of T98B,the germination rate for M1 plants and M2 plants were both significantly suppressed,with a decrease of 22. 35% and 14. 67% respectively. In M2 population,207 candidate mutants were screened. Of which,168 were validated steadily inheritable via a phenotyping test from M3 to M5,accounting for a total mutation frequency of 0. 673%. Furthermore,they were divided into 3 groups regarding leaf group,stem/tiller group and grain/flower group,according to the part that it occurred to mutate on. Each group above included 34,37 and 97 mutants respectively,representing a ratio of 20. 24%,20. 02% and 57. 74%. The common mutants were the ones varied on fertility,leaf color and plant height,and it presented a mutation frequency of 0. 212%,0. 092%and 0. 088% respectively. Moreover,a brief description on the phenotypes for each group was provided. The results indicated that,agronomic mutants in rice could be effectively obtained via gamma-ray irradiation,and several mutants gained here would be severed as useful germ resources for advancing the work on rice genetic improvement.
引文
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[5]邓达胜,陈浩,邓文敏,等.水稻恢复系辐恢838及其衍生系的选育和应用[J].核农学报,2009,23(2):175-179.DENG Dasheng,CHEN Hao,DENG Wenmin,et al.Development of a indica rice restorer line fuhui838 and its derivative lines with strong restoring ability and their utilization[J].Journal of Nuclear Agricultural Sciences,2009,23(2):175-179.
[6]杨仁崔,张书标,黄荣华,等.高秆隐性杂交稻(e-杂交稻)的育种技术[J].中国农业科学,2002,35(3):233-237.YANG Rencui,ZHANG Shubiao,HUANG Ronghua,et al.Breeding technology of eui-hybrids of rice[J].Scientia Agricultura Sinica,2002,35(3):233-237.
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[9]SONG J Y,KIM D S,LEE M C,et al.Physiological characterization of gamma-ray induced salt tolerant rice mutants[J].Australian Journal of Crop Science,2012,6(3):421-429.
[10]ELSHIRBEENY A,MITKEES R.The use of gamma irradiation for inducing high-protein rice[J].Cereal Chemistry,1989,66(2):79-80.
[11]SHU X L,XU J W,WANG Y,et al.Effects of gamma irradiation on starch digestibility of rice with different resistant starch content[J].International Journal of Food Science&Technology,2013,48(1):35-43.
[12]DONG S K,LEE I S,JANG C S,et al.AEC resistant rice mutants induced by gamma-ray irradiation may include both elevated lysine production and increased activity of stress related enzymes[J].Plant Science,2004,167(2):305-316.
[13]HWANG J E,AHN J W,KWON S J,et al.Selection and molecular characterization of a high tocopherol accumulation rice mutant line induced by gamma irradiation[J].Molecular Biology Reports,2014,41(11):7671-7681.
[14]LUOA D,QIAN Q,YIN H F,et al.EUI1,encoding a putative cytochrome P450 monooxygenase,regulates the internodes elongation by modulating GA responses in rice[J].Plant and Cell Physiology,2006,47(2):181-191.
[15]ZHANG H,XU C X,HE Y,et al.Mutation in CSA creates a new photoperiod-sensitive genic male sterile line applicable for hybrid rice seed production[J].Proceedings of the National Academy of Sciences,2013,110(1):76-81.
[16]朱必凤,朱友林,吴成钢,等.对除草剂敏感致死水稻bel基因的RAPD和SCAR分子标记[J].作物学报,2006,32(4):618-624.ZHU Bifeng,ZHU Youlin,WU Chenggang,et al.Identification of RAPD and SCAR markers linked to herbicide susceptible lethality gene(bel)in rice[J].Acta Agronomica Sinica,2006,32(4):618-624.
[17]邓小林,袁定阳,徐荣发.高异交率优质米不育系T98A选育简介[J].杂交水稻,2001,16(4):14.DENG Xiaolin,YUAN Dingyang,XU Rongfa.A brief introduction of T98A,a high outcrossing and good grain quality CMS line[].Hybrid Rice,2001,16(4):14.
[18]谭炎宁,孙学武,袁定阳,等.水稻单叶独立转绿型黄化突变体grc2的鉴定与基因精细定位[J].作物学报,2015,41(6):831-837.TAN Yanning,SUN Xuewu,YUAN Dingyang,et al.Identification and fine mapping of green-revertible chlorina gene grc2in rice(Oryza sativa L.)[J].Acta Agronomica Sinica,2015,41(6):831-837.
[19]TAN Y N,SUN X W,FANG B H,et al.Conversion of a rice CMS maintainer into a photo-or thermo-sensitive genetic male sterile line[J].Molecular Breeding,2018,38(5):56.
[20]SASIKALA R,KALAIYARASI R.Sensitivity of rice varieties to gamma irradiation[J].Electronic Journal of Plant Breeding,2010,1(4):885-889.
[21]郭光荣,易卫平,刘武泉.辐射诱变改造杂交籼稻恢复系的研究[J].中国核科技报告,1995,(S2):480-490.GUO Guangrong,YI Weiping,LIU Wuquan.Improving restorer line of hybrid rice by irradiation[J].China Nuclear Science&Technology Report,1995,(S2):480-490.
[22]STOILOV L,GEORGIEVA M,MANOVA V,et al.Karyotype reconstruction modulates the sensitivity of barley genome to radiation-induced DNA and chromosomal damage[J].Mutagenesis,2013,28(2):153-160.
[23]HEFNER E,PREUSS S B,BRITT A B.Arabidopsis mutant sensitive to gamma radiation include the homologue of the human repair gene ERCC1[J].Journal of Experimental Botany,2003,54(383):669-680.
[24]PARK Y C,KIM J J,KIM D S,et al.Rice ring E3 ligase may negatively regulate gamma-ray response to mediate the degradation of photosynthesis-related proteins[J].Planta,2015,241(5):1119-1129.
[25]GUPTA A,CHRISTENSEN R G,RAYLA A L,et al.An optimized two-finger archive for ZFN-mediated gene targeting[J].Nature Methods,2012,9(6):588-590.
[26]LI T,LIU B,SPALDING M H,et al.High-efficiency TALEN-based gene editing produces disease-resistant rice[J].Nature Biotechnology,2012,30(5):390-392.
[27]ENDO M,MIKAMI M,TOKI S.Multi-gene knockout utilizing off-target mutations of the CRISPR/cas9 system in rice[J].Plant&Cell Physiology,2015,56(1):41-47.
[28]WU J L,WU C,LEI C,et al.Chemical-and irradiation-induced mutants of indica rice IR64 for forward and reverse genetics[J].Plant Molecular Biology,2005,59(1):85-97.
[29]LI G,JAIN R,CHERN M,et al.The sequences of 1 504 mutants in the model rice variety kitaake facilitate rapid functional genomic studies[J].Plant Cell,2017,29(6):1218-1231.
[30]DU H L,YU Y,MA Y F,et al.Sequencing and de novo assembly of a near complete indica rice genome[J].Nature Communications,2017,8:15324.
[31]杨春艳,江玲,沈贝贝,等.水稻品种“南粳35”辐射诱变突变体的鉴定[J].南京农业大学学报,2012,35(3):1-6.YANG Chunyan,JIANG Ling,SHEN Beibei,et al.Identification of mutants from radiated‘Nanjing 35’in rice[J].Journal of Nanjing Agricultural University,2012,35(3):1-6.
[32]SPIELMEYER W,ELLIS M H,CHANDLER P M.Semidwarf(sd-1),“reen revolution”ice,contains a defective gibberellin20-oxidase gene[J].Proceedings of the National Academy of Sciences of the United States of America,2002,99(13):9043-9048.
[33]CHEN Q L,XIE Q J,GAO J,et al.Characterization of rolled and erect leaf 1 in regulating leave morphology in rice[J].Journal of Experimental Botany,2015,66(19):6047-6058.
[34]SAKAMOTO T,KITANO H,FUJIOKA S.An E3 ubiquitin ligase,erect leaf 1,functions in brassinosteroid signaling of rice[J].Plant Signaling&Behavior,2013,8(11):e27117.
[2]邓华凤,舒福北,袁定阳.安农S-1的研究及其利用概况[J].杂交水稻,1999,14(3):1-3.DENG Huafeng,SHU Fubei,YUAN Dingyang.An overview of research and utilization of Annong S-1[J].Hybrid Rice,1999,14(3):1-3.
[3]AHLOOWALIA B S,MALUSZYNSKI M.Induced mutations-a new paradigm in plant breeding[J].Euphytica,2001,118(2):167-173.
[4]RUTGER J N.Applications of induced and spontaneous mutation in rice breeding and genetics[J].Advances in Agronomy,1983,36:383-413.
[5]邓达胜,陈浩,邓文敏,等.水稻恢复系辐恢838及其衍生系的选育和应用[J].核农学报,2009,23(2):175-179.DENG Dasheng,CHEN Hao,DENG Wenmin,et al.Development of a indica rice restorer line fuhui838 and its derivative lines with strong restoring ability and their utilization[J].Journal of Nuclear Agricultural Sciences,2009,23(2):175-179.
[6]杨仁崔,张书标,黄荣华,等.高秆隐性杂交稻(e-杂交稻)的育种技术[J].中国农业科学,2002,35(3):233-237.YANG Rencui,ZHANG Shubiao,HUANG Ronghua,et al.Breeding technology of eui-hybrids of rice[J].Scientia Agricultura Sinica,2002,35(3):233-237.
[7]FUTSUHARA Y,TORIYAMA K,TSUNODA K.Breeding of a new rice variety“reimei”by gamma-ray irradiation[J].Japanese Journal of Breeding,2008,17(2):85-90.
[8]VOTHI M T,NGUYEN T H,PHAN Q M,et al.Applications of gamma rays irradiation and marker assisted selection for improving of bacterial leaf blight resistant rice variety,BT62.1[J].Journal of Agricultural Technology,2015,11(8):2441-2449.
[9]SONG J Y,KIM D S,LEE M C,et al.Physiological characterization of gamma-ray induced salt tolerant rice mutants[J].Australian Journal of Crop Science,2012,6(3):421-429.
[10]ELSHIRBEENY A,MITKEES R.The use of gamma irradiation for inducing high-protein rice[J].Cereal Chemistry,1989,66(2):79-80.
[11]SHU X L,XU J W,WANG Y,et al.Effects of gamma irradiation on starch digestibility of rice with different resistant starch content[J].International Journal of Food Science&Technology,2013,48(1):35-43.
[12]DONG S K,LEE I S,JANG C S,et al.AEC resistant rice mutants induced by gamma-ray irradiation may include both elevated lysine production and increased activity of stress related enzymes[J].Plant Science,2004,167(2):305-316.
[13]HWANG J E,AHN J W,KWON S J,et al.Selection and molecular characterization of a high tocopherol accumulation rice mutant line induced by gamma irradiation[J].Molecular Biology Reports,2014,41(11):7671-7681.
[14]LUOA D,QIAN Q,YIN H F,et al.EUI1,encoding a putative cytochrome P450 monooxygenase,regulates the internodes elongation by modulating GA responses in rice[J].Plant and Cell Physiology,2006,47(2):181-191.
[15]ZHANG H,XU C X,HE Y,et al.Mutation in CSA creates a new photoperiod-sensitive genic male sterile line applicable for hybrid rice seed production[J].Proceedings of the National Academy of Sciences,2013,110(1):76-81.
[16]朱必凤,朱友林,吴成钢,等.对除草剂敏感致死水稻bel基因的RAPD和SCAR分子标记[J].作物学报,2006,32(4):618-624.ZHU Bifeng,ZHU Youlin,WU Chenggang,et al.Identification of RAPD and SCAR markers linked to herbicide susceptible lethality gene(bel)in rice[J].Acta Agronomica Sinica,2006,32(4):618-624.
[17]邓小林,袁定阳,徐荣发.高异交率优质米不育系T98A选育简介[J].杂交水稻,2001,16(4):14.DENG Xiaolin,YUAN Dingyang,XU Rongfa.A brief introduction of T98A,a high outcrossing and good grain quality CMS line[].Hybrid Rice,2001,16(4):14.
[18]谭炎宁,孙学武,袁定阳,等.水稻单叶独立转绿型黄化突变体grc2的鉴定与基因精细定位[J].作物学报,2015,41(6):831-837.TAN Yanning,SUN Xuewu,YUAN Dingyang,et al.Identification and fine mapping of green-revertible chlorina gene grc2in rice(Oryza sativa L.)[J].Acta Agronomica Sinica,2015,41(6):831-837.
[19]TAN Y N,SUN X W,FANG B H,et al.Conversion of a rice CMS maintainer into a photo-or thermo-sensitive genetic male sterile line[J].Molecular Breeding,2018,38(5):56.
[20]SASIKALA R,KALAIYARASI R.Sensitivity of rice varieties to gamma irradiation[J].Electronic Journal of Plant Breeding,2010,1(4):885-889.
[21]郭光荣,易卫平,刘武泉.辐射诱变改造杂交籼稻恢复系的研究[J].中国核科技报告,1995,(S2):480-490.GUO Guangrong,YI Weiping,LIU Wuquan.Improving restorer line of hybrid rice by irradiation[J].China Nuclear Science&Technology Report,1995,(S2):480-490.
[22]STOILOV L,GEORGIEVA M,MANOVA V,et al.Karyotype reconstruction modulates the sensitivity of barley genome to radiation-induced DNA and chromosomal damage[J].Mutagenesis,2013,28(2):153-160.
[23]HEFNER E,PREUSS S B,BRITT A B.Arabidopsis mutant sensitive to gamma radiation include the homologue of the human repair gene ERCC1[J].Journal of Experimental Botany,2003,54(383):669-680.
[24]PARK Y C,KIM J J,KIM D S,et al.Rice ring E3 ligase may negatively regulate gamma-ray response to mediate the degradation of photosynthesis-related proteins[J].Planta,2015,241(5):1119-1129.
[25]GUPTA A,CHRISTENSEN R G,RAYLA A L,et al.An optimized two-finger archive for ZFN-mediated gene targeting[J].Nature Methods,2012,9(6):588-590.
[26]LI T,LIU B,SPALDING M H,et al.High-efficiency TALEN-based gene editing produces disease-resistant rice[J].Nature Biotechnology,2012,30(5):390-392.
[27]ENDO M,MIKAMI M,TOKI S.Multi-gene knockout utilizing off-target mutations of the CRISPR/cas9 system in rice[J].Plant&Cell Physiology,2015,56(1):41-47.
[28]WU J L,WU C,LEI C,et al.Chemical-and irradiation-induced mutants of indica rice IR64 for forward and reverse genetics[J].Plant Molecular Biology,2005,59(1):85-97.
[29]LI G,JAIN R,CHERN M,et al.The sequences of 1 504 mutants in the model rice variety kitaake facilitate rapid functional genomic studies[J].Plant Cell,2017,29(6):1218-1231.
[30]DU H L,YU Y,MA Y F,et al.Sequencing and de novo assembly of a near complete indica rice genome[J].Nature Communications,2017,8:15324.
[31]杨春艳,江玲,沈贝贝,等.水稻品种“南粳35”辐射诱变突变体的鉴定[J].南京农业大学学报,2012,35(3):1-6.YANG Chunyan,JIANG Ling,SHEN Beibei,et al.Identification of mutants from radiated‘Nanjing 35’in rice[J].Journal of Nanjing Agricultural University,2012,35(3):1-6.
[32]SPIELMEYER W,ELLIS M H,CHANDLER P M.Semidwarf(sd-1),“reen revolution”ice,contains a defective gibberellin20-oxidase gene[J].Proceedings of the National Academy of Sciences of the United States of America,2002,99(13):9043-9048.
[33]CHEN Q L,XIE Q J,GAO J,et al.Characterization of rolled and erect leaf 1 in regulating leave morphology in rice[J].Journal of Experimental Botany,2015,66(19):6047-6058.
[34]SAKAMOTO T,KITANO H,FUJIOKA S.An E3 ubiquitin ligase,erect leaf 1,functions in brassinosteroid signaling of rice[J].Plant Signaling&Behavior,2013,8(11):e27117.