油菜素甾醇处理对苹果柱型基因MdCoL和BR相关基因表达的影响
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摘要
柱型苹果(Columnar apple)是选育紧凑型苹果品种的重要资源,其柱型性状由Co基因控制。植物激素被认为是研究苹果柱型性状分子机制的关键点。为进一步分析柱型性状形成和油菜素甾醇(brassinosteroids,BR)之间的关系,本试验用1mg/L BR处理了柱型苹果‘威赛克旭’和转MdCoL基因烟草,用qRT-PCR分析了相关基因表达。结果表明:对照植株(未经BR处理的)柱型苹果和转基因烟草中,BR合成酶基因CYP749A22、85A、734A1、CYP90C和DET2(控制矮化)以及BR信号转导关键酶基因BRI、BZR和BIN2表达量高于处理的;柱型苹果中BAK1基因和724B以及信号转导基因BSK表达量和转基因烟草中的724B、BSK表达量低于处理的。外源BR处理柱型苹果和普通型苹果,MdCoL以及BR合成基因(CYP749A22、CYP90C、DET2和724B)表达量在柱型苹果比普通型苹果低,另一部分合成基因734A1和85A以及信号转导基因BRI、BZR、BIN2、BSK和DIM的表达量比普通型高。外源激素BR处理野生型和转基因烟草后发现,野生型和转基因烟草在节间长度、叶片数、叶绿素含量方面无显著差异;在植株高度上,野生型烟草和CoL-15转基因烟草处理前后无明显差异,未经BR处理的CoL-5转基因烟草在6d之后植株开始生长缓慢,而经BR处理的CoL-5转基因烟草在6d后开始加速生长,说明BR对CoL-5转基因烟草有一定的促进生长作用,而对CoL-15作用不明显。
Columnar apple is an important resource for the selection of compact varieties.The columnar growing habit is controlled by a dominant gene(Co).Hormone regulation is a key point for studying the molecular mechanism of columnar growing habit.In order to further analyze the relationship between the columnar trait and the plant hormones,the expression levels of the hormone-related genes were analyzed using qRT-PCR after exogenous hormone BR(1 mg/L)treatment on McIntosh(columnar type)apple and MdCoLtransformed tobacco plants.The results were as follows:The expression of BR syntheses genes CYP749 A,85 A,734 A1,CYP90 Cand DET2(dwarfing genes),the signal genes BRI,BZR,and BIN2 were higher in the untreated columnar apple and the MdCoLtransformed tobacco plants than those in treated ones.In columnar apples,the synthetic gene BAK1 and 724 B,the signal transduction gene BSK were all lower;In transgenic tobacco,the 724 B,the signal transduction gene BSK were all lower than those in treated ones.The expression of MdCoL and BR synthetic genes(CYP749 A22,CYP90 C,DET2 and724 B)of columnar apples after exogenous BR treatment of columnar and wild type apples was lower than that of the WT,and another part of the synthetic genes 734 A1,85 Aand signal transduction genes BRI,BZR,BIN2,BSKand DIMexpression is higher than the WT.There was no significant difference in internode length,leaf number and chlorophyll content between the wild-type and transgenic MdCoLtobacco plants treated with hormone BR(1 mg/L).There was also no significant differnce in the plant height between the wild-type tobacco and CoL-15 T3 MdCoLtransformed tobacco plants treated with BR(1 mg/L).However,the CoL-5 T3 MdCoLtransformed tobacco plants began to grow slowly 6 days after the spray of achole while the BR-treated CoL-5 began to accelerate 6 days after the spray of BR(1 mg/L),indicating BR would have a certain growth-promoting effect on the CoL-5 transgenic plants and no obvious effect on the CoL-15 transgenic plants.
Columnar apple is an important resource for the selection of compact varieties.The columnar growing habit is controlled by a dominant gene(Co).Hormone regulation is a key point for studying the molecular mechanism of columnar growing habit.In order to further analyze the relationship between the columnar trait and the plant hormones,the expression levels of the hormone-related genes were analyzed using qRT-PCR after exogenous hormone BR(1 mg/L)treatment on McIntosh(columnar type)apple and MdCoLtransformed tobacco plants.The results were as follows:The expression of BR syntheses genes CYP749 A,85 A,734 A1,CYP90 Cand DET2(dwarfing genes),the signal genes BRI,BZR,and BIN2 were higher in the untreated columnar apple and the MdCoLtransformed tobacco plants than those in treated ones.In columnar apples,the synthetic gene BAK1 and 724 B,the signal transduction gene BSK were all lower;In transgenic tobacco,the 724 B,the signal transduction gene BSK were all lower than those in treated ones.The expression of MdCoL and BR synthetic genes(CYP749 A22,CYP90 C,DET2 and724 B)of columnar apples after exogenous BR treatment of columnar and wild type apples was lower than that of the WT,and another part of the synthetic genes 734 A1,85 Aand signal transduction genes BRI,BZR,BIN2,BSKand DIMexpression is higher than the WT.There was no significant difference in internode length,leaf number and chlorophyll content between the wild-type and transgenic MdCoLtobacco plants treated with hormone BR(1 mg/L).There was also no significant differnce in the plant height between the wild-type tobacco and CoL-15 T3 MdCoLtransformed tobacco plants treated with BR(1 mg/L).However,the CoL-5 T3 MdCoLtransformed tobacco plants began to grow slowly 6 days after the spray of achole while the BR-treated CoL-5 began to accelerate 6 days after the spray of BR(1 mg/L),indicating BR would have a certain growth-promoting effect on the CoL-5 transgenic plants and no obvious effect on the CoL-15 transgenic plants.
引文
[1]Lapins KO.Inheritance of compact growth type in apple[J].JAm Soc Hortic Sci,1976,101(2):133-135
[2]Lapins KO.Segregation of compact growth types in certain apple seedling progenies[J].Can J Plant Sci,1969,49(6):765-768
[3]Tobutt KR.Breeding columnar apple trees at east malling[J].Acta Hort,1985,159(159):63-68
[4]Tobutt KR.Combining apetalous parthenocarpy with columnar growth habit in apple[J].Euphytica,1994,77(1):51-54
[5]Zhang YG,Dai HY.Comparison of photosynthetic and morphological characteristics,and micro-structure of roots and shoots,between columnar apple and standard apple trees of hybrid seedlings[J].Phyton,2011,80(2):119-125
[6]Sarwar M,Skirvin RM,Kushad M,et al.Selecting dwarf apple(Malus x domestica Borkh.)Trees in vitro:multiple cytokinin tolerance expressed among three strains of‘‘McIntosh’’that differ in their growth habit under field conditions[J].Plant Cell Tiss Org,1998,54(2):71-76
[7]Liu A,Eaton GW.Comparative leaf anatomy of two standard and two compact apple mutants[J].Can J Plant Sci,1970,50(6):733-735
[8]Gelvonauskis B,Brazaityte A,Sasnauskas A,et al.Morphological and physiological characteristics of columnar apple trees[J].Scientific Works of the Lithuanian Institute of Horticulture and Lithuanian University of Agriculture,2006,25(5):350-356
[9]Moriya S,Iwanami H,Kotoda N,et al.Development of a marker-assisted selection system for columnar growth habit in apple breeding[J].Journal of the Japanese Society for Horticultural Science,2009,78(3):279-287
[10]Moriya S,Okada K,Haji T,et al.Fine mapping of Co,agene controlling columnar growth habit located on apple(Malus×domestica)linkage group 10[J].Plant Breeding,2012,131(5):641-647
[11]Meulenbroek J,Verhaegh J,Janse J.Inheritance studies with columnar type trees[J].Acta Hort 1998,1(484):255-258
[12]Tian YK,Wang CH,Zhang JS,et al.Mapping Co,agene controlling the columnar phenotype of apple,with molecular markers[J].Euphytica,2005,145(1):181-188
[13]王彩虹,王倩,戴洪义,等.与柱型苹果基因Co紧密连锁额分子标记的筛选.农业生物技术学报[J].2001,9(2):187-190
[14]王彩虹,王倩,戴洪义,等.苹果柱型基因Co的一个AFLP标记的SCAR转换.园艺学报[J].2001,29(2):100-104
[15]Zhang YG,Zhu J,Dai HY.Characterization of transcriptional differences between columnar and standard apple trees using RNA-Seq[J].Plant Molecular Biology Reporter,2010,30(4):957-965
[16]Kawai Y,Ono E,Mizutani M.Evolution and diversity of the 2-oxoglutarate-dependent dioxygenase superfamily in plant[J].Plant Journal,2014,78(2):328-343
[17]Okada K,Wada M,Moriya S,et al.Expression of putative dioxygenase gene adjacent to an insertion mutation is involved in the short internodes of columnar apple(Malus×domestica)[J].Journal of Plant Research,2016,129(6):1109-1126
[18]Otto D,Petersen R,Brauksiepe B,et al.The columnar mutation(“Co”gene)of apple(Malus×domestica)is accociated with an intergration of a Gypsy-like retrotransposon[J].Molecular breeding,2014,33(4):863-880
[19]Bai TH,Zhu YD,Ferna′ndez-Ferna′ndez F,et al.Fine genetic mapping of the Co locus controlling columnar growth habit in apple[J].Molecular Genetics and Genomics,2012,287(5):437-450
[20]Shimada Y,Goda H,Nakamura A,et al.Organ-specific expression of brassinosteroid-biosynthetic genes and distribution of endogenous brassinosteroids in Arabidopsis[J].Plant physiol,2003,131(1):287-297
[21]Li H,Jiang L,Youn JH,et al.A comprehensive genetic study reveals a crucial role of CYP90D2/D2in regulating plant architecture in rice(Oryza sativa)[J].New Phytol,2013,200(4):1076-1088
[22]Kim TW,Guan S,Sun Y,et al.Brassinosteroid signal transduction from cell-surface receptor kinases to nuclear transcription factors[J].Nat Cell Biol,2009,11(10):1254-1260
[23]Ohnishi T,Nomura T,Watanabe B,et al.Tomato cytochrome P450 CYP734A7functions in brassinosteroid catabolism[J].Phytochem,2006,67(17):1895-1906
[24]Takahashi T,Gasch A,Nishizawa N,et al.The IMINUTOgene of Arabidopsis is involved in regulating cell elongation[J].Genes Dev,1995,1(9):97-107
[25]Wang H,Nagegowda DA,Rawat R,et al.Overexpression of Brassica juncea wildtype and mutant HMG-CoA synthase 1in Arabidopsis upregulates genes in sterol biosynthesis and enhances sterol production and stress tolerance[J].Plant Biotechnol J,2011,10(3):31-42
[26]Holender CA,Dardick C.Molecular basis of angiosperm tree architecture[J]New Phytol,2015,206(2):541-56
[27]Petersen R,Krost C.Tracting a key player in the regulation of plant architecture:the columnar growth habit of apple trees(Malus x domestica)[J].Planta,2013,238(2):1-22
[2]Lapins KO.Segregation of compact growth types in certain apple seedling progenies[J].Can J Plant Sci,1969,49(6):765-768
[3]Tobutt KR.Breeding columnar apple trees at east malling[J].Acta Hort,1985,159(159):63-68
[4]Tobutt KR.Combining apetalous parthenocarpy with columnar growth habit in apple[J].Euphytica,1994,77(1):51-54
[5]Zhang YG,Dai HY.Comparison of photosynthetic and morphological characteristics,and micro-structure of roots and shoots,between columnar apple and standard apple trees of hybrid seedlings[J].Phyton,2011,80(2):119-125
[6]Sarwar M,Skirvin RM,Kushad M,et al.Selecting dwarf apple(Malus x domestica Borkh.)Trees in vitro:multiple cytokinin tolerance expressed among three strains of‘‘McIntosh’’that differ in their growth habit under field conditions[J].Plant Cell Tiss Org,1998,54(2):71-76
[7]Liu A,Eaton GW.Comparative leaf anatomy of two standard and two compact apple mutants[J].Can J Plant Sci,1970,50(6):733-735
[8]Gelvonauskis B,Brazaityte A,Sasnauskas A,et al.Morphological and physiological characteristics of columnar apple trees[J].Scientific Works of the Lithuanian Institute of Horticulture and Lithuanian University of Agriculture,2006,25(5):350-356
[9]Moriya S,Iwanami H,Kotoda N,et al.Development of a marker-assisted selection system for columnar growth habit in apple breeding[J].Journal of the Japanese Society for Horticultural Science,2009,78(3):279-287
[10]Moriya S,Okada K,Haji T,et al.Fine mapping of Co,agene controlling columnar growth habit located on apple(Malus×domestica)linkage group 10[J].Plant Breeding,2012,131(5):641-647
[11]Meulenbroek J,Verhaegh J,Janse J.Inheritance studies with columnar type trees[J].Acta Hort 1998,1(484):255-258
[12]Tian YK,Wang CH,Zhang JS,et al.Mapping Co,agene controlling the columnar phenotype of apple,with molecular markers[J].Euphytica,2005,145(1):181-188
[13]王彩虹,王倩,戴洪义,等.与柱型苹果基因Co紧密连锁额分子标记的筛选.农业生物技术学报[J].2001,9(2):187-190
[14]王彩虹,王倩,戴洪义,等.苹果柱型基因Co的一个AFLP标记的SCAR转换.园艺学报[J].2001,29(2):100-104
[15]Zhang YG,Zhu J,Dai HY.Characterization of transcriptional differences between columnar and standard apple trees using RNA-Seq[J].Plant Molecular Biology Reporter,2010,30(4):957-965
[16]Kawai Y,Ono E,Mizutani M.Evolution and diversity of the 2-oxoglutarate-dependent dioxygenase superfamily in plant[J].Plant Journal,2014,78(2):328-343
[17]Okada K,Wada M,Moriya S,et al.Expression of putative dioxygenase gene adjacent to an insertion mutation is involved in the short internodes of columnar apple(Malus×domestica)[J].Journal of Plant Research,2016,129(6):1109-1126
[18]Otto D,Petersen R,Brauksiepe B,et al.The columnar mutation(“Co”gene)of apple(Malus×domestica)is accociated with an intergration of a Gypsy-like retrotransposon[J].Molecular breeding,2014,33(4):863-880
[19]Bai TH,Zhu YD,Ferna′ndez-Ferna′ndez F,et al.Fine genetic mapping of the Co locus controlling columnar growth habit in apple[J].Molecular Genetics and Genomics,2012,287(5):437-450
[20]Shimada Y,Goda H,Nakamura A,et al.Organ-specific expression of brassinosteroid-biosynthetic genes and distribution of endogenous brassinosteroids in Arabidopsis[J].Plant physiol,2003,131(1):287-297
[21]Li H,Jiang L,Youn JH,et al.A comprehensive genetic study reveals a crucial role of CYP90D2/D2in regulating plant architecture in rice(Oryza sativa)[J].New Phytol,2013,200(4):1076-1088
[22]Kim TW,Guan S,Sun Y,et al.Brassinosteroid signal transduction from cell-surface receptor kinases to nuclear transcription factors[J].Nat Cell Biol,2009,11(10):1254-1260
[23]Ohnishi T,Nomura T,Watanabe B,et al.Tomato cytochrome P450 CYP734A7functions in brassinosteroid catabolism[J].Phytochem,2006,67(17):1895-1906
[24]Takahashi T,Gasch A,Nishizawa N,et al.The IMINUTOgene of Arabidopsis is involved in regulating cell elongation[J].Genes Dev,1995,1(9):97-107
[25]Wang H,Nagegowda DA,Rawat R,et al.Overexpression of Brassica juncea wildtype and mutant HMG-CoA synthase 1in Arabidopsis upregulates genes in sterol biosynthesis and enhances sterol production and stress tolerance[J].Plant Biotechnol J,2011,10(3):31-42
[26]Holender CA,Dardick C.Molecular basis of angiosperm tree architecture[J]New Phytol,2015,206(2):541-56
[27]Petersen R,Krost C.Tracting a key player in the regulation of plant architecture:the columnar growth habit of apple trees(Malus x domestica)[J].Planta,2013,238(2):1-22