香蕉BADH基因序列及表达特性分析
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摘要
甜菜碱醛脱氢酶(BADH)是植物体内甜菜碱合成过程中的关键酶,对植物抵御逆境具有重要作用。为研究香蕉BADH基因序列及表达特性,本研究利用香蕉基因数据库对香蕉BADH基因序列及蛋白序列进行生物学信息分析,采用qTR-PCR技术对香蕉幼苗中BADH基因进行表达分析。结果表明:MaBADH基因属单克隆基因,含有15个外显子,14个内含子;顺式调控元件预测显示在MaBADH启动子区域存在15种激素和逆境胁迫相关的顺式调控元件;蛋白结构域和功能分析有一个甜菜碱醛脱氢酶结构域具有氧化还原酶活性;亚细胞定位在叶绿体中,多重序列比对发现其具有N-端信号肽和C-端信号肽两种信号肽;系统进化树显示其与水稻、玉米、菠萝亲缘关系最近。基因表达分析发现MaBADH具有组织特异表达特性,在叶片中相对表达量最高,假茎次之,根最低;此外,茉莉酸甲酯能够诱导香蕉幼苗根和叶中MaBADH差异表达,其中对叶片的诱导能力更强。本研究为激素信号调控甜菜碱分子保护机制和利用基因工程提高香蕉抗逆性提供了参考依据。
Betaine aldehyde dehydrogenase(BADH) is a key enzyme in the process of betaine synthesis in plants,which plays an important role in stress resistance of plants. To study the sequence and expression characteristics of banana BADH gene, the gene sequence and protein sequence of BADH in banana from banana genome database were analyzed by bioinformatics, and the expression analysis of BADH in banana seedlings was revealed by q TR-PCR technology in this study. The results showed that the MaBADH gene is a monoclonal gene containing 15 exons and 14 introns. Cis-regulatory element prediction showed that there were 15 species hormone and stress-related cis-regulatory elements in MaBADH promoter regions. Protein domain and functional analysis showed that MaBADH has a betaine aldehyde dehydrogenase domain with oxidoreductase activity. This gene may be localized in the chloroplast and has two signal peptides: an N-terminal signal peptide and a C-terminal signal peptide. The phylogenetic tree indicated that BADH of banana was closer to rice, corn and pineapple in the homologous relationship. Expression analysis showed that MaBADH has tissue specific expression properties, and the relative expession of the middle leaves of most MaBADH was the highest, followed by the pseudostems and the roots. In addition,methyl jasmonate can induce differential expression of MaBADH in roots and leaves of banana seedlings, and the ability to induce leaves is stronger. This study provides a reference for hormone signaling to regulate the molecular protection mechanism of betaine and banana resistance improvement by genetic engineering.
Betaine aldehyde dehydrogenase(BADH) is a key enzyme in the process of betaine synthesis in plants,which plays an important role in stress resistance of plants. To study the sequence and expression characteristics of banana BADH gene, the gene sequence and protein sequence of BADH in banana from banana genome database were analyzed by bioinformatics, and the expression analysis of BADH in banana seedlings was revealed by q TR-PCR technology in this study. The results showed that the MaBADH gene is a monoclonal gene containing 15 exons and 14 introns. Cis-regulatory element prediction showed that there were 15 species hormone and stress-related cis-regulatory elements in MaBADH promoter regions. Protein domain and functional analysis showed that MaBADH has a betaine aldehyde dehydrogenase domain with oxidoreductase activity. This gene may be localized in the chloroplast and has two signal peptides: an N-terminal signal peptide and a C-terminal signal peptide. The phylogenetic tree indicated that BADH of banana was closer to rice, corn and pineapple in the homologous relationship. Expression analysis showed that MaBADH has tissue specific expression properties, and the relative expession of the middle leaves of most MaBADH was the highest, followed by the pseudostems and the roots. In addition,methyl jasmonate can induce differential expression of MaBADH in roots and leaves of banana seedlings, and the ability to induce leaves is stronger. This study provides a reference for hormone signaling to regulate the molecular protection mechanism of betaine and banana resistance improvement by genetic engineering.
引文
Arnold K.,B ordoli L.,Kopp J.,and Schwede T.,2006,Th swiss-model workspace:a web-based environment for protein structure homology modelling,Bioinformatics,22(2):195-201
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Cong H.Q.,Qi Y.Y.,Zhang Z.W.,Chen S.B.,and Qiao F.,2015,Sequence analysis of EIN3 gene and expression characteristics induced by ethylene and methyl jasmonate in Cassava(Manihot esculenta Crantz),Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),13(12):2705-2712(丛汉卿,齐尧尧,张振文,陈松笔,乔飞,2015,木薯EIN3基因序列分析及乙烯和茉莉酸甲酯诱导表达特性,分子植物育种,13(12):2705-2712)
Gasteiger E.,Gattiker A.,Hoogland C.,Ivanyi I.,Appel R.D.,and Bairoch A.,2003,ExPASy:the proteomics server for in-depth protein knowledge and analysis,Nucleic Acids Res.,31(13):3784-3788
Hanson A.D.,May A.M.,Rebecca G.,Janet B.,Gene C.J.,and David R.,1985,Grumet R.,betaine synthesis in chenopodes:localization in choroplasts,Proc.Natl.Acad.Sci.USA,82(1):3678-3695
Hu B.,Jin J.P.,Guo A.Y.,Zhang H.,Luo J.C.,and Gao G.,2014,GSDS 2.0:an upgraded gene feature visualization server,Bioinformatics,31(8):1296-1297
Hu L.Z.,Wang J.S.,Zhao J.H.,Wang Q.X.,Han J.Y.,and Ji X.E.,2017,Identification and comparative analysis of the TIFYfamily genes in tomato and potato,Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),15(4):1192-1203(胡利宗,王俊生,赵锦慧,王秋霞,韩娇月,纪秀娥,2017,番茄与马铃薯TIFY家族基因的鉴定与比较分析,分子植物育种,15(4):1192-1203)
Jiang X.M.,Huang M.R.,and Wang M.X.,2002,A review on betaine-biosynthesized pathway and genetic engineering in plants,Zhongguo Shengwu Gongcheng Zhazhi(Journal of Chinese Biotechnology),22(4):49-56(江香梅,黄敏仁,王明庥,2002,植物甜菜碱合成途径及基因工程研究进展,中国生物工程杂志,22(4):49-56)
Johansson K.,El-Ahmad M.,Ramaswamy S.,and Hjelmqvist L.,1998,Structure of betaine aldehyde dehydrogenase at 2.1 a resolution,Protein Sci.,7(10):2106-2117
Jones D.T.,1999,Protein secondary structure prediction based on position-specific scoring matrices,J.Mol.Biol.,292(2):195-202
Krogh A.,Larsson B.,Von Heijne G.,and Sonnhammer E.L.,2001,Predicting transmembrane protein topology with a hidden Markov model:application to complete genomes,J.Mol.Biol.,305(3):567-580
Lescot M.,Déhais P.,Thijs G.,Marchal K.,Moreau Y.,Van dePeer Y.,RouzéP.,and Rombauts S.,2002,PlantCARE:a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences,Nucleic Acids Res.,30(1):325-327
Li Y.H.,Wang W.,Yang X.H.,and Zou Q.,2005,Expression of BADH and betaine content in wheat cultivars with different drought resistance under drought stress,Zuowu Xuebao(Acta Agronomica Sinica),31(4):425-430(李永华,王玮,杨兴洪,邹琦,2005,干旱胁迫下不同抗旱性小麦BADH表达及甜菜碱含量的变化,作物学报,31(4):425-430)
Liang Z.,and Luo A.L.,1995,Betaine and betaine synthetase,Zhiwu Shenglixue Tongxun(Plant Physiology Communications),32(1):1-8(梁峥,骆爱玲,1995,甜菜碱和甜菜碱合成酶,植物生理学通讯,31(1):1-8)
Liu Z.L.,and Dai S.L.,2004,Studies on betaine aldeh yde deh ydrogenase(BADH)gene in plants,Xibei Nonglin Keji Daxue Xuebao(Journal of Northwest A&F University),32(3):104-112(刘振林,戴思兰,2004,植物甜菜碱醛脱氢酶基因研究进展,西北农林科技大学学报,32(3):104-112)
Liu Z.L.,Chao H.W.,Xia X.L.,Yi W.L.,and Dai S.L.,2009,Cloning of BADH gene cDNA of chamomile and its expression under salt stress,Wuhan Zhiwuxue Yanjiu(Wuhan Botanical Research),27(1):1-7(刘振林,曹华雯,夏新莉,尹伟伦,戴思兰,2009,甘菊BADH基因cDNA的克隆及在盐胁迫下的表达,武汉植物学研究,27(1):1-7)
Long Y.L.,Xu Z.J.,Zhu B.B.,Jiang X.F.,and Zhang W.,2016,Sequence analysis of ClTPS1 gene in watermelon and its expression characteristic after treated with MeJA,Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),14(12):3299-3307(龙娅丽,徐子健,朱白婢,江雪飞,张文,2016,茉莉酸甲酯诱导西瓜ClTPS1基因的表达特性及其生物学信息分析,分子植物育种,14(12):3299-3307)
Lu P.,Tian Y.S.,Wang M.X.,Li S.,and Zhao J.Y.,2013,Cloning and expression analysis of a key enzyme gene NCED of abscisic acid biosynthesis,Zhiwu Yichuan Zhiyuan Xuebao(Journal of Plant Genetic Resources),14(2):303-310(陆平,田跃胜,王名雪,李杉,赵静雅,2013,枸杞脱落酸生物合成关键酶基因NCED的克隆及表达分析,植物遗传资源学报,14(2):303-310)
Lv L.M.,Zhang S.,Luo J.Y.,Wang C.Y.,Wang L.,Zhu X.Z.,Zhang L.J.,Li C.H.,and Cui J.J.,2018,Effects of beet armyworm feeding and methyl jasmonate treatment on transcription levels of key genes involved in the biosynthesis pathway of jasmonic acid and terpene synthase in cotton,Zhiwu Shengli Xuebao(Plant Physiology Journal),54(4):636-644(吕丽敏,张帅,雒珺瑜,王春义,王丽,朱香镇,张利娟,李春花,崔金杰,2018,甜菜夜蛾和茉莉酸甲酯处理对棉花茉莉酸合成途径关键基因及萜类合酶基因表达的影响,植物生理学报,54(4):636-644)
McCue K.F.,and Hanson A.D.,1992,Salt-inducible betaine aldehyde dehydrogenase from sugar beet:cDNA cloning and expression,Plant Mol.Biol.,18(1):1-11
Nakamura T.,Yokota S.,Muramoto Y.,Tsutsui K.,Oguri Y.,Fukui K.,and Takabe T.,1997,Expression of a betaine aldehyde dehydrogenase gene in rice,a glycinebetaine nonaccumulator,and possible localization of its protein in peroxisomes,Plant J.,11(5):1115-1120
Niu J.S.,Liu J.,Ni Y.J.,and Yin J.,2011,Induction of jasmonic acid on PR-1,PR-2,PR-5 and Ta-JA2 gene expression and resistance to wheat powdery mildew,Zhiwu Bingli Xuebao(Acta Phytopathologica Sinica),41(3):270-277(牛吉山,刘靖,倪永静,尹钧,2011,茉莉酸对PR-1,PR-2,PR-5和Ta-JA2基因表达以及小麦白粉病抗性的诱导,植物病理学报,41(3):270-277)
Nomura M.,Takabe T.,and Sugiyama T.,1998,Transgenicall produced glycinebetatine protects ribulose 1,5-bisphodphate carboxylase/oxygenase from inactivation in Synechococcus sp.PCC7942 under salt stress,Plant Cell Physiol.,39(4):425-432
Qin D.,Zhao C.L.,Zheng C.Z.,and Wang P.W.,2015,Drought tolerance of transgenic soybean with BADH gene,Zhongguo Youliao Zuowu Xuebao(Chinese Journal of Oil Crop Sciences),37(6):752-758(秦迪,赵翠兰,郑成忠,王丕武,2015,转BADH基因大豆耐旱性分析,中国油料作物学报,37(6):752-758)
Rajendra B.,Jonathan D.,and Jones G.,2009,Role of plant hormones in plant defence responses,Plant Mol.Biol.,69(4):473-488
Wang B.,and Li W.M.,2001,Advances in signal transduction pathways of plant hormones,Fujian Nnongye Daxue Xuebao(Journal of Fujian Agricultural University),30(4):433-438(汪斌,李维明,2001,植物激素的信号转导系统研究进展,福建农业大学学报,30(4):433-438)
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Wang W.C.,Zhou S.Y.,Ji F.S.,Li Y.Y.,and Li X.G.,2017,The relative expression of CaM and Ca2+-ATPase genes in banana seedlings under salt stress by real-time fluorescence quantitative assay,Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),15(5):1745-1751(王文昌,周双云,乔飞,吉福桑,李元元,李新国,2017,实时荧光定量检测盐胁迫下香蕉幼苗CaM和Ca2+-ATPase基因的相对表达量,分子植物育种,15(5):1745-1751)
Wang X.L.,Du J.Z.,Hao Y.S.,Zhang L.J.,Zhao X.M.,Wang Y.X.,and Sun Y.,2014,Transformation of BADH gene into maize and salt tolerence of transgenic plant,Zuowu Xuebao(Acta Agronomica Sinica),40(11):1973-1979(王小丽,杜建中,郝曜山,张丽君,赵欣梅,王亦学,孙毅,2014,转BADH基因玉米植株的获得及其耐盐性分析,作物学报,40(11):1973-1979)
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Wood A.J.,Saneoka H.,Rhodes D.,Joly R.J.,and Goldsbrough P.B.,1996,Betaine aldehyde dehydrogenase in sorghum,P-lant Physiol.,110(4):1301-1308
Xu Z.J.,Sun M.L.,Long Y.L.,Qiao F.,Dang X.M.,and Jiang X.F.,2017,Sequence analysis of BADH gene and expression characteristics induced by ethylene and methyl jasmonate in watermelon,Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),15(4):1218-1225(徐子健,孙梦利,龙娅丽,乔飞,党选民,江雪飞,2017,西瓜BADH基因序列分析及其乙烯和茉莉酸甲酯诱导表达特性,分子植物育种,15(4):1218-1225)
Zhang C.H.,Gong P.J.,Wei R.,Li S.X.,Zhang X.T.,Yu Y.H.,and Wang Y.J.,2013,The metacaspase gene family of Vitis vinifera L:characterization and differential expression during ovule abortion in stenospermocarpic seedless grapes,G-ene,528(2):267-276
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Bateman J.B.,Evans G.F.,Brown P.R.,Gabriel C.,and Grant E.H.,1992,Dielectric properties of the system bovine albu min:urea:betaine in aqueous solution,Phys.Med.Biol.,37(1):175-182
Bouizgarne B.,El-Maarouf-Bouteau H.,Frankart C.,Reboutier D.,Madiona K.,Pennarun A.M.,Monestiez M.,Trouverie J.,Amiar Z.,Briand J.,Braul M.,Rona J.P.,Ouhdouch Y.,El adrami I.,and Bouteau F.,2006,Early physiological responses of Arabidopsis thaliana cells to fusaric acid:toxic and signaling effects,New Phytol.,169(1):209-218
Cong H.Q.,Qi Y.Y.,Zhang Z.W.,Chen S.B.,and Qiao F.,2015,Sequence analysis of EIN3 gene and expression characteristics induced by ethylene and methyl jasmonate in Cassava(Manihot esculenta Crantz),Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),13(12):2705-2712(丛汉卿,齐尧尧,张振文,陈松笔,乔飞,2015,木薯EIN3基因序列分析及乙烯和茉莉酸甲酯诱导表达特性,分子植物育种,13(12):2705-2712)
Gasteiger E.,Gattiker A.,Hoogland C.,Ivanyi I.,Appel R.D.,and Bairoch A.,2003,ExPASy:the proteomics server for in-depth protein knowledge and analysis,Nucleic Acids Res.,31(13):3784-3788
Hanson A.D.,May A.M.,Rebecca G.,Janet B.,Gene C.J.,and David R.,1985,Grumet R.,betaine synthesis in chenopodes:localization in choroplasts,Proc.Natl.Acad.Sci.USA,82(1):3678-3695
Hu B.,Jin J.P.,Guo A.Y.,Zhang H.,Luo J.C.,and Gao G.,2014,GSDS 2.0:an upgraded gene feature visualization server,Bioinformatics,31(8):1296-1297
Hu L.Z.,Wang J.S.,Zhao J.H.,Wang Q.X.,Han J.Y.,and Ji X.E.,2017,Identification and comparative analysis of the TIFYfamily genes in tomato and potato,Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),15(4):1192-1203(胡利宗,王俊生,赵锦慧,王秋霞,韩娇月,纪秀娥,2017,番茄与马铃薯TIFY家族基因的鉴定与比较分析,分子植物育种,15(4):1192-1203)
Jiang X.M.,Huang M.R.,and Wang M.X.,2002,A review on betaine-biosynthesized pathway and genetic engineering in plants,Zhongguo Shengwu Gongcheng Zhazhi(Journal of Chinese Biotechnology),22(4):49-56(江香梅,黄敏仁,王明庥,2002,植物甜菜碱合成途径及基因工程研究进展,中国生物工程杂志,22(4):49-56)
Johansson K.,El-Ahmad M.,Ramaswamy S.,and Hjelmqvist L.,1998,Structure of betaine aldehyde dehydrogenase at 2.1 a resolution,Protein Sci.,7(10):2106-2117
Jones D.T.,1999,Protein secondary structure prediction based on position-specific scoring matrices,J.Mol.Biol.,292(2):195-202
Krogh A.,Larsson B.,Von Heijne G.,and Sonnhammer E.L.,2001,Predicting transmembrane protein topology with a hidden Markov model:application to complete genomes,J.Mol.Biol.,305(3):567-580
Lescot M.,Déhais P.,Thijs G.,Marchal K.,Moreau Y.,Van dePeer Y.,RouzéP.,and Rombauts S.,2002,PlantCARE:a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences,Nucleic Acids Res.,30(1):325-327
Li Y.H.,Wang W.,Yang X.H.,and Zou Q.,2005,Expression of BADH and betaine content in wheat cultivars with different drought resistance under drought stress,Zuowu Xuebao(Acta Agronomica Sinica),31(4):425-430(李永华,王玮,杨兴洪,邹琦,2005,干旱胁迫下不同抗旱性小麦BADH表达及甜菜碱含量的变化,作物学报,31(4):425-430)
Liang Z.,and Luo A.L.,1995,Betaine and betaine synthetase,Zhiwu Shenglixue Tongxun(Plant Physiology Communications),32(1):1-8(梁峥,骆爱玲,1995,甜菜碱和甜菜碱合成酶,植物生理学通讯,31(1):1-8)
Liu Z.L.,and Dai S.L.,2004,Studies on betaine aldeh yde deh ydrogenase(BADH)gene in plants,Xibei Nonglin Keji Daxue Xuebao(Journal of Northwest A&F University),32(3):104-112(刘振林,戴思兰,2004,植物甜菜碱醛脱氢酶基因研究进展,西北农林科技大学学报,32(3):104-112)
Liu Z.L.,Chao H.W.,Xia X.L.,Yi W.L.,and Dai S.L.,2009,Cloning of BADH gene cDNA of chamomile and its expression under salt stress,Wuhan Zhiwuxue Yanjiu(Wuhan Botanical Research),27(1):1-7(刘振林,曹华雯,夏新莉,尹伟伦,戴思兰,2009,甘菊BADH基因cDNA的克隆及在盐胁迫下的表达,武汉植物学研究,27(1):1-7)
Long Y.L.,Xu Z.J.,Zhu B.B.,Jiang X.F.,and Zhang W.,2016,Sequence analysis of ClTPS1 gene in watermelon and its expression characteristic after treated with MeJA,Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),14(12):3299-3307(龙娅丽,徐子健,朱白婢,江雪飞,张文,2016,茉莉酸甲酯诱导西瓜ClTPS1基因的表达特性及其生物学信息分析,分子植物育种,14(12):3299-3307)
Lu P.,Tian Y.S.,Wang M.X.,Li S.,and Zhao J.Y.,2013,Cloning and expression analysis of a key enzyme gene NCED of abscisic acid biosynthesis,Zhiwu Yichuan Zhiyuan Xuebao(Journal of Plant Genetic Resources),14(2):303-310(陆平,田跃胜,王名雪,李杉,赵静雅,2013,枸杞脱落酸生物合成关键酶基因NCED的克隆及表达分析,植物遗传资源学报,14(2):303-310)
Lv L.M.,Zhang S.,Luo J.Y.,Wang C.Y.,Wang L.,Zhu X.Z.,Zhang L.J.,Li C.H.,and Cui J.J.,2018,Effects of beet armyworm feeding and methyl jasmonate treatment on transcription levels of key genes involved in the biosynthesis pathway of jasmonic acid and terpene synthase in cotton,Zhiwu Shengli Xuebao(Plant Physiology Journal),54(4):636-644(吕丽敏,张帅,雒珺瑜,王春义,王丽,朱香镇,张利娟,李春花,崔金杰,2018,甜菜夜蛾和茉莉酸甲酯处理对棉花茉莉酸合成途径关键基因及萜类合酶基因表达的影响,植物生理学报,54(4):636-644)
McCue K.F.,and Hanson A.D.,1992,Salt-inducible betaine aldehyde dehydrogenase from sugar beet:cDNA cloning and expression,Plant Mol.Biol.,18(1):1-11
Nakamura T.,Yokota S.,Muramoto Y.,Tsutsui K.,Oguri Y.,Fukui K.,and Takabe T.,1997,Expression of a betaine aldehyde dehydrogenase gene in rice,a glycinebetaine nonaccumulator,and possible localization of its protein in peroxisomes,Plant J.,11(5):1115-1120
Niu J.S.,Liu J.,Ni Y.J.,and Yin J.,2011,Induction of jasmonic acid on PR-1,PR-2,PR-5 and Ta-JA2 gene expression and resistance to wheat powdery mildew,Zhiwu Bingli Xuebao(Acta Phytopathologica Sinica),41(3):270-277(牛吉山,刘靖,倪永静,尹钧,2011,茉莉酸对PR-1,PR-2,PR-5和Ta-JA2基因表达以及小麦白粉病抗性的诱导,植物病理学报,41(3):270-277)
Nomura M.,Takabe T.,and Sugiyama T.,1998,Transgenicall produced glycinebetatine protects ribulose 1,5-bisphodphate carboxylase/oxygenase from inactivation in Synechococcus sp.PCC7942 under salt stress,Plant Cell Physiol.,39(4):425-432
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