水稻抗稻瘟病基因多重PCR检测技术建立及应用
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摘要
利用水稻稻瘟病抗性基因建立2套多重PCR体系,体系Ⅰ、Ⅱ分别同时检测稻瘟病抗性基因Pikh与Pi1、Pita与Pib,并用这2套体系对114份杂交后代水稻材料中抗病基因进行检测。结果表明:114份材料中含Pita抗性基因的有81份,含Pib的有106份,含Pikh的有35份,含Pi1的有3份。穗颈瘟接种鉴定结果显示,114份材料中人工接种鉴定表现为高抗的有2份,抗病的有27份,中抗的有32份,感病的有33份,高感的有20份;Pita、Pikh基因与穗颈瘟的抗性呈显著正相关,相关系数分别为0.297、0.239;Pib、Pi1基因与穗颈瘟的抗性相关不显著。
In this study,two multiple PCR systems were established to detect the blast resistance genes.The PCR system I was used to detect the resistance alleles on Pi1 and Pikh,and the PCR system II was used to detect the resistance alleles on Pitaand Pib.Both systems were used to detect blast-resistant genes in 114 hybrid rice materials and the results showed that there were 81 lines contained Pita,106 lines contained Pib,35 lines contained Pikhand 3 lines contained Pi1.Artificial inoculation of blast in rice ear was conducted to demonstrate the resistance of these 114 materials to blast.Among these 114 materials,2 lines manifested highly resistance,27 lines manifested resistance,32 lines manifested moderate resistance,33 lines showed susceptible,20 lines showed highly susceptible.Further correlation analyses between blast resistance and resistant genes revealed that Pita and Pikh were positively correlated with blast resistance of rice(P<0.05),and the correlation coefficients were 0.297 and 0.239,respectively.Pib and Pi1 had no significant correlation with the resistance of panicle blast.
In this study,two multiple PCR systems were established to detect the blast resistance genes.The PCR system I was used to detect the resistance alleles on Pi1 and Pikh,and the PCR system II was used to detect the resistance alleles on Pitaand Pib.Both systems were used to detect blast-resistant genes in 114 hybrid rice materials and the results showed that there were 81 lines contained Pita,106 lines contained Pib,35 lines contained Pikhand 3 lines contained Pi1.Artificial inoculation of blast in rice ear was conducted to demonstrate the resistance of these 114 materials to blast.Among these 114 materials,2 lines manifested highly resistance,27 lines manifested resistance,32 lines manifested moderate resistance,33 lines showed susceptible,20 lines showed highly susceptible.Further correlation analyses between blast resistance and resistant genes revealed that Pita and Pikh were positively correlated with blast resistance of rice(P<0.05),and the correlation coefficients were 0.297 and 0.239,respectively.Pib and Pi1 had no significant correlation with the resistance of panicle blast.
引文
[1]潘争艳,邱福林,吕桂兰,等.辽星系列水稻品种稻瘟病抗性基因鉴定与抗性分析[J].江苏农业科学,2017,45(7):87-90.
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[3]阎勇,马增凤,秦钢,等.华南常用籼稻亲本稻瘟病抗性评价及抗性基因鉴定[J].南方农业学报,2017,48(4):587-593.
[4]王慧,陈金节,严志,等.利用分子标记辅助选择改良不育系03S的稻瘟病抗性[J].扬州大学学报(农业与生命科学版),2015,36(4):74-78.
[5]余玲,戴正元,吴云雨,等.利用分子标记辅助选择提高江苏粳稻稻瘟病抗性[J].扬州大学学报(农业与生命科学版),2014,35(4):57-62.
[6]鄂志国,张丽靖,焦桂,等.稻瘟病抗性基因的鉴定及利用进展[J].中国水稻科学,2008,22(5):533-540.
[7]范方军,王芳权,刘永峰,等.Pi-b、Pi-ta、Pikm和Pi54对水稻穗颈瘟的抗性评价[J].华北农学报,2014,29(3):221-226.
[8]王军,杨杰,陈志德,等.利用分子标记辅助选择聚合水稻抗病基因Pita、Pi-b和Stv-bi[J].作物学报,2011,37(6):975-981.
[9]姚姝,刘燕清,张亚东,等.水稻抗稻瘟病基因Pi-ta和Pi-b多重PCR体系的构建与应用[J].Agricultural Science&Technology,2015,40(10):2225-2230.
[10]MA W,ZHANG W,GALE K R.Multiplex-PCR typing of high molecular weight glutenin alleles in wheat[J].Euphytica,2003,134(1):51-60.
[11]NAKAMURA T,VRINTEN P,SAITO M,et al.Rapid classification of partial waxy wheats using PCR-based markers[J].Genome,2002,45(6):1150.
[12]罗楚平,倪磊,陈志谊,等.水稻稻瘟病接种技术及2009年江苏省区试品种抗性鉴定[J].江苏农业科学,2009(6):178-179.
[13]邱福林,王和和,陈洁,等.用于水稻突变体大量筛选的DNA微量快速提取法[J].中国水稻科学,2006,20(3):329-332.
[14]刘洋,徐培洲,张红宇,等.水稻抗稻瘟病Pib基因的分子标记辅助选择与应用[J].中国农业科学,2008,41(1):9-14.
[15]STEFANO C,JIA Y.Sequence variation at the rice blast resistance gene Pi-kmlocus:implications for the development of allele specific markers[J].Plant Science,2010,178(6):523-530.
[16]RAMKUMAR G,SRINIVASARAO K,MOHAN K M,et al.Development and validation of functional marker targeting an InDel in the major rice blast disease resistance gene Pi54(Pikh)[J].Molecular Breeding,2011,27(1):129-135.
[17]JIA Y L,WANG Z H,SINGH P.Development of dominant rice blast Pi-ta resistance gene markers[J].Crop Science,2003,42(6):2145-2149.
[18]FJELLSTROM R,CONAWAYBORMANS C A,MCCLUNG A M,et al.Development of DNA markers suitable for marker assisted selection of three Pi genes conferring resistance to multiple Pyricularia grisea pathotypes[J].Crop Science,2004,44(5):1790-1798.
[19]ZHAI C,LIN F,DONG Z,et al.The isolation and characterization of Pik,a rice blast resistance gene which emerged after rice domestication[J].New Phytologist,2011,189(1):321.
[20]陈涛,张亚东,朱镇,等.Pi-b和Pi-ta基因在江苏省粳稻中的分布以及与穗颈瘟抗性的关系[J].江苏农业学报,2016,32(1):1-8.
[21]王军,宫丹妮,杨杰,等.江苏省粳稻品种抗稻瘟病基因型与穗颈瘟抗性分析[J].江苏农业学报,2016,32(2):250-256.
[2]李刚,王健,曹奎荣,等.江苏水稻主导品种的稻瘟病抗性及抗性基因遗传多样性[J].湖南农业大学学报(自然科学版),2017,43(3):238-243.
[3]阎勇,马增凤,秦钢,等.华南常用籼稻亲本稻瘟病抗性评价及抗性基因鉴定[J].南方农业学报,2017,48(4):587-593.
[4]王慧,陈金节,严志,等.利用分子标记辅助选择改良不育系03S的稻瘟病抗性[J].扬州大学学报(农业与生命科学版),2015,36(4):74-78.
[5]余玲,戴正元,吴云雨,等.利用分子标记辅助选择提高江苏粳稻稻瘟病抗性[J].扬州大学学报(农业与生命科学版),2014,35(4):57-62.
[6]鄂志国,张丽靖,焦桂,等.稻瘟病抗性基因的鉴定及利用进展[J].中国水稻科学,2008,22(5):533-540.
[7]范方军,王芳权,刘永峰,等.Pi-b、Pi-ta、Pikm和Pi54对水稻穗颈瘟的抗性评价[J].华北农学报,2014,29(3):221-226.
[8]王军,杨杰,陈志德,等.利用分子标记辅助选择聚合水稻抗病基因Pita、Pi-b和Stv-bi[J].作物学报,2011,37(6):975-981.
[9]姚姝,刘燕清,张亚东,等.水稻抗稻瘟病基因Pi-ta和Pi-b多重PCR体系的构建与应用[J].Agricultural Science&Technology,2015,40(10):2225-2230.
[10]MA W,ZHANG W,GALE K R.Multiplex-PCR typing of high molecular weight glutenin alleles in wheat[J].Euphytica,2003,134(1):51-60.
[11]NAKAMURA T,VRINTEN P,SAITO M,et al.Rapid classification of partial waxy wheats using PCR-based markers[J].Genome,2002,45(6):1150.
[12]罗楚平,倪磊,陈志谊,等.水稻稻瘟病接种技术及2009年江苏省区试品种抗性鉴定[J].江苏农业科学,2009(6):178-179.
[13]邱福林,王和和,陈洁,等.用于水稻突变体大量筛选的DNA微量快速提取法[J].中国水稻科学,2006,20(3):329-332.
[14]刘洋,徐培洲,张红宇,等.水稻抗稻瘟病Pib基因的分子标记辅助选择与应用[J].中国农业科学,2008,41(1):9-14.
[15]STEFANO C,JIA Y.Sequence variation at the rice blast resistance gene Pi-kmlocus:implications for the development of allele specific markers[J].Plant Science,2010,178(6):523-530.
[16]RAMKUMAR G,SRINIVASARAO K,MOHAN K M,et al.Development and validation of functional marker targeting an InDel in the major rice blast disease resistance gene Pi54(Pikh)[J].Molecular Breeding,2011,27(1):129-135.
[17]JIA Y L,WANG Z H,SINGH P.Development of dominant rice blast Pi-ta resistance gene markers[J].Crop Science,2003,42(6):2145-2149.
[18]FJELLSTROM R,CONAWAYBORMANS C A,MCCLUNG A M,et al.Development of DNA markers suitable for marker assisted selection of three Pi genes conferring resistance to multiple Pyricularia grisea pathotypes[J].Crop Science,2004,44(5):1790-1798.
[19]ZHAI C,LIN F,DONG Z,et al.The isolation and characterization of Pik,a rice blast resistance gene which emerged after rice domestication[J].New Phytologist,2011,189(1):321.
[20]陈涛,张亚东,朱镇,等.Pi-b和Pi-ta基因在江苏省粳稻中的分布以及与穗颈瘟抗性的关系[J].江苏农业学报,2016,32(1):1-8.
[21]王军,宫丹妮,杨杰,等.江苏省粳稻品种抗稻瘟病基因型与穗颈瘟抗性分析[J].江苏农业学报,2016,32(2):250-256.