功能性雄性不育茄子SmLOX基因的克隆与表达分析
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摘要
本研究以茄子(Solanum melongena L.)功能性雄性不育系F13-1-7、T4和正常可育茄子142为材料,基于茄子基因组数据库,利用PCR方法克隆获得SmLOX基因的cDNA全长序列。结果表明,SmLOX基因的cDNA全长分别为2 557 bp,开放阅读框长度为2 508 bp,编码836个氨基酸,蛋白分子量为94.52 kD,理论等电点p I为5.3。进化树分析表明该基因与马铃薯、番茄的亲缘关系较近。亚细胞定位显示,Sm LOX蛋白N端均不含信号肽。定量分析发现,不育系中SmLOX的表达量都低于可育系,表明这个基因参与调控花药开裂过程。这为深入研究茄子功能性雄性不育和茄科植物花药开裂分子调控机制提供了一定的科学依据。
Based on the information of transcriptome database derived from eggplant(Solanum melongena L.),the cDNA sequences of SmLOX was cloned from male sterile(F13-1-7, T4) and fertile(142) material. Sequence analysis showed that the full length of c DNAs was 2 557 bp, with 2 508 bp open reading frames(ORF). That cDNAs encoded 836 amino acids. The molecular weight of the protein was 94.52 kD and the pI was 5.3. The results of phylogenetic analysis showed that SmLOX gene had a close relationship with Solanum tuberosum and Solanum lycop-ersicum, respectively. Subcellular localization analysis indicated that SmLOX protein did not contain signal peptides at the N-terminus. The analysis of the expression of the gene showed that the expression level of SmLOX in the male sterile lines was lower than those in the fertile lines, indicating that the gene was involved in the regulation of the anther dehiscence process. These results provided the basis for in-depth study of functional male sterility and the mechanism of anther dehiscence.
Based on the information of transcriptome database derived from eggplant(Solanum melongena L.),the cDNA sequences of SmLOX was cloned from male sterile(F13-1-7, T4) and fertile(142) material. Sequence analysis showed that the full length of c DNAs was 2 557 bp, with 2 508 bp open reading frames(ORF). That cDNAs encoded 836 amino acids. The molecular weight of the protein was 94.52 kD and the pI was 5.3. The results of phylogenetic analysis showed that SmLOX gene had a close relationship with Solanum tuberosum and Solanum lycop-ersicum, respectively. Subcellular localization analysis indicated that SmLOX protein did not contain signal peptides at the N-terminus. The analysis of the expression of the gene showed that the expression level of SmLOX in the male sterile lines was lower than those in the fertile lines, indicating that the gene was involved in the regulation of the anther dehiscence process. These results provided the basis for in-depth study of functional male sterility and the mechanism of anther dehiscence.
引文
Bannenberg G.,Martinez M.,Hamberg M.,and Castresana C.,2009,Diversity of the enzymatic activity in the lipoxygenase gene family of Arabidopsis thaliana,Lipids,44(2):85-95
Burow G.B.,Gardner H.W.,and Keller N.P.,2000,A peanut seed lipoxygenase respo nsive to Aspergillus colonization,Plant Mol.Biol.,42(5):689-701
Caldelari D.,Wang G.,Farmer E.E.,and Dong X.,2011,Arabidopsis lox3 lox4 double mutants are male sterile and defective in global proliferative arrest,Plant Mol.Biol.,75(1-2):25-33
Feys B.J.F.,Benedetti C.E.,Penfold C.N.,and Turner J.G.,1994,Arabidopsis mutants selected for resistance to the phytotoxin cronatine are male-sterile,insensitive to methyl jasmonate,and resistant to a bacterial pathogen,Plant Cell,6(5):751-759
Fonseca S.,Chico J.M.,and Solano R.,2009,The jasmonate pathway:the ligand,the receptor and the core signalling module,Curr.Opin.Plant Biol.,12(5):539-547
Glauser G.,Dubugnon L.,Mousavi S.A.,Rudaz S.,Wolfender J.L.,and Farmer E.E.,2009,Velocity estimates for signal propagation leading to s ystemic jasmonic acid accumulation in wounded Arabidopsis,J.Biol.Chem.,284(50):34506-34513
Katsir L.,Chung H.S.,Koo A.J.K.,and Howe G.A.,2008,Jasmonate signaling:a conserved mechanism of hormone sensing,Curr.Opin.Plant Biol.,11(4):428-435
Li X.J.,Liu H.B.,Lin X.Q.,Wu Z.D.,Xu C.H.,and Liu X.L.,2015,In silico cloning and bioinformatics analysis of KNOXgene in sugarcane(Sckn1),Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),34(1):136-142(李旭娟,刘洪博,林秀琴,吴转娣,徐超华,刘新龙,2015,甘蔗KNOX基因(Sckn1)的电子克隆及生物信息学分析,基因组学与应用生物学,34(1):136-142)
McConn M.,and Browse J.,1996,The critical requirement for linolenic acid is pollen development,not photosynthesis,in an Arabidopsis mutant,Plant Cell,8(3):403-416
Needleman P.,Turk J.,Jakschik B.A.,Morrison A.R.,and Lefkowith J.B.,1986,Arachidonic-acid metabolism,Annu.Rev.Biochem.,55(4):69-102
Peng Y.J.,Shih C.F.,Yang J.Y.,Tan C.M.,Hsu W.H.,Huang Y.P.,Liao P.C.,and Yang C.H.,2013,A RING-type E3 ligase controls anther dehiscence by activating the jasmonate biosynthetic pathway gene DEFECTIVE IN ANTHER DE-HISCENCE1 in Arabidopsis,Plant J.,74(2):310-327
Scott R.J.,Spielman M.,and Dickinson H.G.,2004,Stamen structure and function,Plant Cell,16(S):46-60
Seltmann M.A.,Stingl N.E.,Lautenschlaeger J.K.,Krischke M.,Mueller M.J.,and Berger S.,2010,Differential impact of lipoxygenase 2 and jasmonates on natural and stress-induced senescence in Arabidopsis,Plant Physiol.,152(4):1940-1950
Song S.S.,Qi T.C.,Huang H.,Ren Q.C.,Wu D.W.,Chang C.Q.,Peng W.,Liu Y.L.,Peng J.R.,and Xie D.X.,2011,The jasmonate-ZIM domain proteins interact with the R2R3-MYBtranscription factors MYB21 and MYB24 to affect jasmonateregulated stamen development in Arabidopsis,Plant Cell,23(3):1000-1013
Tomchick D.R.,Phan P.,Cymborowski M.,Minor W.,and Holman T.R.,2001,Structural and functional characterization of second-coordination sphere mutants of soybean lipoxpgenase-1,Biochemistry,40(25):7509-7517
Wasternack C.,and Hause B.,2013,Jasmonates:biosynthesis,perception,signal transduction and action in plant stress response,growth and development,An update to the 2007review in annals of botany,Ann.Bot.,111(6):1021-1058
Zhou P.F.,Jia R.R.,Chen S.C.,Xu L.Z.,Peng A.H.,Lai T.G.,Li Q.,Chen M.,Bai X.J.,Zou X.P.,and He Y.R.,2017,Cloning and expression analysis of four citrus WRKY genes responding to Xanthomon asaxonopodis pv.Citri,Yuanyi Xuebao(Acta Horticulturae Sinica)44(3):452-462(周鹏飞,贾瑞瑞,陈善春,许兰珍,彭爱红,雷天刚,李强,陈敏,白晓晶,邹修平,何永睿,2017,柑橘4个WRKY转录因子基因的克隆及其响应柑橘溃疡病菌侵染的表达分析,园艺学报,44(3):452-462)
Burow G.B.,Gardner H.W.,and Keller N.P.,2000,A peanut seed lipoxygenase respo nsive to Aspergillus colonization,Plant Mol.Biol.,42(5):689-701
Caldelari D.,Wang G.,Farmer E.E.,and Dong X.,2011,Arabidopsis lox3 lox4 double mutants are male sterile and defective in global proliferative arrest,Plant Mol.Biol.,75(1-2):25-33
Feys B.J.F.,Benedetti C.E.,Penfold C.N.,and Turner J.G.,1994,Arabidopsis mutants selected for resistance to the phytotoxin cronatine are male-sterile,insensitive to methyl jasmonate,and resistant to a bacterial pathogen,Plant Cell,6(5):751-759
Fonseca S.,Chico J.M.,and Solano R.,2009,The jasmonate pathway:the ligand,the receptor and the core signalling module,Curr.Opin.Plant Biol.,12(5):539-547
Glauser G.,Dubugnon L.,Mousavi S.A.,Rudaz S.,Wolfender J.L.,and Farmer E.E.,2009,Velocity estimates for signal propagation leading to s ystemic jasmonic acid accumulation in wounded Arabidopsis,J.Biol.Chem.,284(50):34506-34513
Katsir L.,Chung H.S.,Koo A.J.K.,and Howe G.A.,2008,Jasmonate signaling:a conserved mechanism of hormone sensing,Curr.Opin.Plant Biol.,11(4):428-435
Li X.J.,Liu H.B.,Lin X.Q.,Wu Z.D.,Xu C.H.,and Liu X.L.,2015,In silico cloning and bioinformatics analysis of KNOXgene in sugarcane(Sckn1),Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),34(1):136-142(李旭娟,刘洪博,林秀琴,吴转娣,徐超华,刘新龙,2015,甘蔗KNOX基因(Sckn1)的电子克隆及生物信息学分析,基因组学与应用生物学,34(1):136-142)
McConn M.,and Browse J.,1996,The critical requirement for linolenic acid is pollen development,not photosynthesis,in an Arabidopsis mutant,Plant Cell,8(3):403-416
Needleman P.,Turk J.,Jakschik B.A.,Morrison A.R.,and Lefkowith J.B.,1986,Arachidonic-acid metabolism,Annu.Rev.Biochem.,55(4):69-102
Peng Y.J.,Shih C.F.,Yang J.Y.,Tan C.M.,Hsu W.H.,Huang Y.P.,Liao P.C.,and Yang C.H.,2013,A RING-type E3 ligase controls anther dehiscence by activating the jasmonate biosynthetic pathway gene DEFECTIVE IN ANTHER DE-HISCENCE1 in Arabidopsis,Plant J.,74(2):310-327
Scott R.J.,Spielman M.,and Dickinson H.G.,2004,Stamen structure and function,Plant Cell,16(S):46-60
Seltmann M.A.,Stingl N.E.,Lautenschlaeger J.K.,Krischke M.,Mueller M.J.,and Berger S.,2010,Differential impact of lipoxygenase 2 and jasmonates on natural and stress-induced senescence in Arabidopsis,Plant Physiol.,152(4):1940-1950
Song S.S.,Qi T.C.,Huang H.,Ren Q.C.,Wu D.W.,Chang C.Q.,Peng W.,Liu Y.L.,Peng J.R.,and Xie D.X.,2011,The jasmonate-ZIM domain proteins interact with the R2R3-MYBtranscription factors MYB21 and MYB24 to affect jasmonateregulated stamen development in Arabidopsis,Plant Cell,23(3):1000-1013
Tomchick D.R.,Phan P.,Cymborowski M.,Minor W.,and Holman T.R.,2001,Structural and functional characterization of second-coordination sphere mutants of soybean lipoxpgenase-1,Biochemistry,40(25):7509-7517
Wasternack C.,and Hause B.,2013,Jasmonates:biosynthesis,perception,signal transduction and action in plant stress response,growth and development,An update to the 2007review in annals of botany,Ann.Bot.,111(6):1021-1058
Zhou P.F.,Jia R.R.,Chen S.C.,Xu L.Z.,Peng A.H.,Lai T.G.,Li Q.,Chen M.,Bai X.J.,Zou X.P.,and He Y.R.,2017,Cloning and expression analysis of four citrus WRKY genes responding to Xanthomon asaxonopodis pv.Citri,Yuanyi Xuebao(Acta Horticulturae Sinica)44(3):452-462(周鹏飞,贾瑞瑞,陈善春,许兰珍,彭爱红,雷天刚,李强,陈敏,白晓晶,邹修平,何永睿,2017,柑橘4个WRKY转录因子基因的克隆及其响应柑橘溃疡病菌侵染的表达分析,园艺学报,44(3):452-462)