菠萝AcWRKY71-like基因的分离与克隆分析
|
摘要
WRKY转录因子在植物应对胁迫和生长发育过程中都具有重要的调节功能。乙烯诱导菠萝开花为生产常规栽培技术,然而其分子机制尚未揭晓。为了探索菠萝AcWRKY71-like基因的生物学功能,本研究克隆了该基因,并对AcWRKY71-like蛋白进行生物信息学分析,最后检测了AcWRKY71-like基因对外源乙烯的响应特性。结果显示:该基因全长为1 221 bp,开放阅读框(ORF)为909 bp;AcWRKY71-like蛋白属于亲水性蛋白,包含28个磷酸化位点且不包含跨膜结构域和信号肽;而且外源乙烯能够较快并稳定地诱导AcWRKY71-like基因的表达。基于此,我们最终构建了35S::AcWRKY71-like表达载体。本研究为探索菠萝AcWRKY71-like基因的功能以及乙烯诱导菠萝开花分子机制提供了参考依据。
WRKY transcription factors play an important regulatory role in plant response to stress and growth.Ethylene induced pineapple flowering is a conventional cultivation technique. However, its molecular mechanism has not yet been revealed. In order to explore the biological function of AcWRKY71-like gene in pineapple, the AcWRKY71-like gene was cloned, and the AcWRKY71-like protein was analyzed by bioinformatics. Finally, the response characteristics of AcWRKY71-like gene to exogenous ethylene were detected. The results showed tha the full length of the gene was 1 212 bp, and the open reading frame(ORF) was 909 bp. AcWRKY71-like protein was the hydrophilic protein, including 28 phosphorylation sites without transmembrane domain and signal peptide.Exogenous ethylene could induce the expression of AcWRKY71-like gene more quickly and stably. Based on this,we finally constructed 35 S::AcWRKY71-like expression vector. This study provides a reference for exploring the function of AcWRKY71-like gene in pineapple and the molecular mechanism of ethylene-induced flowering in pineapple.
WRKY transcription factors play an important regulatory role in plant response to stress and growth.Ethylene induced pineapple flowering is a conventional cultivation technique. However, its molecular mechanism has not yet been revealed. In order to explore the biological function of AcWRKY71-like gene in pineapple, the AcWRKY71-like gene was cloned, and the AcWRKY71-like protein was analyzed by bioinformatics. Finally, the response characteristics of AcWRKY71-like gene to exogenous ethylene were detected. The results showed tha the full length of the gene was 1 212 bp, and the open reading frame(ORF) was 909 bp. AcWRKY71-like protein was the hydrophilic protein, including 28 phosphorylation sites without transmembrane domain and signal peptide.Exogenous ethylene could induce the expression of AcWRKY71-like gene more quickly and stably. Based on this,we finally constructed 35 S::AcWRKY71-like expression vector. This study provides a reference for exploring the function of AcWRKY71-like gene in pineapple and the molecular mechanism of ethylene-induced flowering in pineapple.
引文
Bleecker A.B.,and Patt erson S.E.,1997,Last exit:senescence,abscission,and meristem arrest in Arabidopsis,Plant Cell,9(7):1169-1179
Das S.C.,Prakash J.,Suresh C.P.,Das A.,and Bhattacharjee T.,2011,Pineapple cultivationin hilly Tripura with year around production:improving livelihood opportunities in rural areas of Tripura,Acta Horticulturae,902:291-298
Espinosa M.E.á.,Moreira R.O.,Lima A.A.,Ságio S.A.,Barreto H.G.,Luiz S.L.P.,Abreu C.E.A.,Yanes-Paz E.,Ruíz Y.C.,González-Olmedo J.L.,and Chalfun-Júnior A.,2017,Early histological,hormonal,and molecular changes during pineapple(Ananas comosus(L.)Merrill)artificial flowering induction,J.Plant Physiol.,209:11-19
Eulgem T.,Rushton P.J.,Robatzek S.,and Somssich I.E.,2000,The WRKY superfamily of plant transcription factors,Trends Plant Sci.,5(5):199-206
Lei M.,Li Z.Y.,Wang Z.,and Xu L.,2016,Cloning and tissue specific expression analysis of TERMINAL FLOWER 1 in Aechemia fasciata,Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),14(1):45-50(雷明,李志英,王之,徐立,2016,蜻蜓凤梨TERMINAL FLOWER 1基因的克隆及组织表达分析,分子植物育种,14(1):45-50)
Liang R.Y.,Chen X.Y.,Sun S.H.,Sun H.X.,and Dun W.T.,2017,Bioinformatics analysis of TFAM gene encoding protein in sheep,Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),36(2):575-581(梁瑞圆,陈晓勇,孙少华,孙洪新,敦伟涛,2017,绵羊TFAM基因编码蛋白生物信息学分析,基因组学与应用生物学,32(6):575-581)
Long F.,Li S.P.,and Li M.F.,2013,Bioinformatics analysis of ALAD gene in seven plants,Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),32(6):802-814(龙芳,李绍鹏,李茂富,2013,7种植物ALAD基因的生物信息学分析,基因组学与应用生物学,32(6):802-814)
Maruthasalam S.,Shiu L.Y.,Loganathan M.,Lien W.C.,Liu Y.L.,Sun C.M.,Yu C.W.,Hung S.H.,Ko Y.,and Lin C.H.,2010,Forced flowering of pineapple(Ananas comosus cv.Tainon 17)in response to cold stress,ethephon and calcium carbide with or without activated charcoal,Plant Growth Regul.,60(2):83-90
Trusov Y.,and Botella J.R.,2006,Silencing of the ACC synthase gene ACACS2 causes delayed flowering in pineapple[Ananas comosus(L.)Merr.],J.Exp.Bot.,57(14):3953-3960
Wang J.B.,Li Z.Y.,Lei M.,Fu Y.L.,Zhao J.J.,Ao M.F.,and Xu L.,2017,Integrated DNA methylome and transcriptome analysis reveals the ethylene-induced flowering pathway genes in pineapple,Sci.Rep.,7(1):17167
Wang N.,Zhang Z.B.,and Huang F.Z.,2014,Advance on the Role of WRKY transcription factors in abiotic stress response,Henongxue Bao(Journal of Nuclear Agricultural Sciences),28(10):1819-1827(王娜,张振葆,黄凤珠,2014,WRKY转录因子参与植物非生物胁迫应答的研究进展,核农学报,28(10):1819-1827)
Xiao X.,Zhang J.M.,and Feng J.,2016,The amplification and targeting vector construction of a polyketide synthases gene of Fonsecaea monophora,Zhongguo Zhenjunxue Zazhi(Chinese Journal of Mycology),11(2):70-74(肖星,张军民,冯姣,2016,Fonsecaea monophora多聚酮合酶基因的扩增及敲除载体的构建,中国真菌学杂志,11(2):70-74)
Xie T.,Chen C.J.,Li C.H.,Liu J.R.,Liu C.Y.,and He Y.H.,2018,Genome-wide investigation of WRKY gene family in pineapple:evolution and expression profiles during development and stress,BMC Genomics,19(1):490
Xie Z.W.,Wang L.J.,and Chen J.Y.,2016,Studies on WRKYtranscription factors and their biological functions in plants,Zhongguo Nongye Keji Daobao(Journal of Agricultural Science and Techology),18(3):46-54(谢政文,王连军,陈锦洋,2016,植物WRKY转录因子及其生物学功能研究进展,中国农业科技导报,18(3):46-54)
Das S.C.,Prakash J.,Suresh C.P.,Das A.,and Bhattacharjee T.,2011,Pineapple cultivationin hilly Tripura with year around production:improving livelihood opportunities in rural areas of Tripura,Acta Horticulturae,902:291-298
Espinosa M.E.á.,Moreira R.O.,Lima A.A.,Ságio S.A.,Barreto H.G.,Luiz S.L.P.,Abreu C.E.A.,Yanes-Paz E.,Ruíz Y.C.,González-Olmedo J.L.,and Chalfun-Júnior A.,2017,Early histological,hormonal,and molecular changes during pineapple(Ananas comosus(L.)Merrill)artificial flowering induction,J.Plant Physiol.,209:11-19
Eulgem T.,Rushton P.J.,Robatzek S.,and Somssich I.E.,2000,The WRKY superfamily of plant transcription factors,Trends Plant Sci.,5(5):199-206
Lei M.,Li Z.Y.,Wang Z.,and Xu L.,2016,Cloning and tissue specific expression analysis of TERMINAL FLOWER 1 in Aechemia fasciata,Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),14(1):45-50(雷明,李志英,王之,徐立,2016,蜻蜓凤梨TERMINAL FLOWER 1基因的克隆及组织表达分析,分子植物育种,14(1):45-50)
Liang R.Y.,Chen X.Y.,Sun S.H.,Sun H.X.,and Dun W.T.,2017,Bioinformatics analysis of TFAM gene encoding protein in sheep,Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),36(2):575-581(梁瑞圆,陈晓勇,孙少华,孙洪新,敦伟涛,2017,绵羊TFAM基因编码蛋白生物信息学分析,基因组学与应用生物学,32(6):575-581)
Long F.,Li S.P.,and Li M.F.,2013,Bioinformatics analysis of ALAD gene in seven plants,Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),32(6):802-814(龙芳,李绍鹏,李茂富,2013,7种植物ALAD基因的生物信息学分析,基因组学与应用生物学,32(6):802-814)
Maruthasalam S.,Shiu L.Y.,Loganathan M.,Lien W.C.,Liu Y.L.,Sun C.M.,Yu C.W.,Hung S.H.,Ko Y.,and Lin C.H.,2010,Forced flowering of pineapple(Ananas comosus cv.Tainon 17)in response to cold stress,ethephon and calcium carbide with or without activated charcoal,Plant Growth Regul.,60(2):83-90
Trusov Y.,and Botella J.R.,2006,Silencing of the ACC synthase gene ACACS2 causes delayed flowering in pineapple[Ananas comosus(L.)Merr.],J.Exp.Bot.,57(14):3953-3960
Wang J.B.,Li Z.Y.,Lei M.,Fu Y.L.,Zhao J.J.,Ao M.F.,and Xu L.,2017,Integrated DNA methylome and transcriptome analysis reveals the ethylene-induced flowering pathway genes in pineapple,Sci.Rep.,7(1):17167
Wang N.,Zhang Z.B.,and Huang F.Z.,2014,Advance on the Role of WRKY transcription factors in abiotic stress response,Henongxue Bao(Journal of Nuclear Agricultural Sciences),28(10):1819-1827(王娜,张振葆,黄凤珠,2014,WRKY转录因子参与植物非生物胁迫应答的研究进展,核农学报,28(10):1819-1827)
Xiao X.,Zhang J.M.,and Feng J.,2016,The amplification and targeting vector construction of a polyketide synthases gene of Fonsecaea monophora,Zhongguo Zhenjunxue Zazhi(Chinese Journal of Mycology),11(2):70-74(肖星,张军民,冯姣,2016,Fonsecaea monophora多聚酮合酶基因的扩增及敲除载体的构建,中国真菌学杂志,11(2):70-74)
Xie T.,Chen C.J.,Li C.H.,Liu J.R.,Liu C.Y.,and He Y.H.,2018,Genome-wide investigation of WRKY gene family in pineapple:evolution and expression profiles during development and stress,BMC Genomics,19(1):490
Xie Z.W.,Wang L.J.,and Chen J.Y.,2016,Studies on WRKYtranscription factors and their biological functions in plants,Zhongguo Nongye Keji Daobao(Journal of Agricultural Science and Techology),18(3):46-54(谢政文,王连军,陈锦洋,2016,植物WRKY转录因子及其生物学功能研究进展,中国农业科技导报,18(3):46-54)