RNA-seq挖掘四倍体小麦紫籽主效基因
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摘要
紫色籽粒四倍体小麦籽粒中富含花青素,是优良的遗传学标记,但其分子遗传机理尚不清楚。本研究利用RNA-seq技术比较四倍体小麦紫粒品种‘深紫1号’和白粒品种‘高原3号’果皮转录组,挖掘控制紫粒性状的主效基因。结果表明紫色和白色果皮中共检测到95 404个Unigenes。21 373个基因表达差异显著,其中3 891个基因在紫色果皮表达上调,12 482个基因表达下调。差异表达基因主要与次生代谢途径相关,其中黄酮类和花青素合成代谢通路差异最为明显。除F3'H外,与花青素合成相关的结构基因在紫色果皮中的表达水平均高于白色,说明花青素合成代谢与四倍体小麦的紫粒性状相关。调控花青素合成代谢的MYC转录因子Unigene25029_All位于2AL染色体上,在紫色果皮中表达水平比白色果皮高出4.78倍,与普通小麦中控制紫粒性状的TaMYC1 (pp3)同源,这个基因很有可能就是四倍体小麦紫色果皮性状的候选基因。本研究结果筛选到四倍体小麦中紫粒性状的候选基因,有助于了解小麦紫色果皮性状的起源及其分子机理的解析。
The purple seeds of tetraploid wheat are rich in anthocyanin, which is an excellent genetic marker, but its molecular genetic mechanism is not clear. In this study, the transcriptome of its pericarp was compared with'Purple1' and 'Gaoyuan3' for digging the key genes responsible for controling the purple grain character. The results showed that 95 404 unigenes were detected in purple and white pericarp. There were 21 373 Unigenes significantly differentially in expression, including 3 891 up-regulated in purple pericarp and 17 482 down-regulated in white percicrap. The differentially expressed unigenes were mainly involved in secondary metabolic pathways, among them, the flavonoid biosynthesis pathway was the most represented in metabolic pathways. Most structural genes related to anthocyanin biosynthesis had higher expression level in purple than white except F3'H, which implied that anthocyanin biosynthesis was related to purple grain traits of tetraploid wheat. The MYC transcription factor Unigene25029_All was located on chromosome 2 AL, and its expression level in purple pericarp was 4.78 times higher than that in white pericarp, and homologous to TaMYC1(pp3), which controls purple pericarp in common wheat. This gene was likely to be a candidate for purple pericarp traits in teraploid wheat. In this article, candidate gene for purple pericarp traits in teraploid wheat was screened, which might be helpful to understand the origin and molecular mechanism of purple pericarp traits in wheat.
The purple seeds of tetraploid wheat are rich in anthocyanin, which is an excellent genetic marker, but its molecular genetic mechanism is not clear. In this study, the transcriptome of its pericarp was compared with'Purple1' and 'Gaoyuan3' for digging the key genes responsible for controling the purple grain character. The results showed that 95 404 unigenes were detected in purple and white pericarp. There were 21 373 Unigenes significantly differentially in expression, including 3 891 up-regulated in purple pericarp and 17 482 down-regulated in white percicrap. The differentially expressed unigenes were mainly involved in secondary metabolic pathways, among them, the flavonoid biosynthesis pathway was the most represented in metabolic pathways. Most structural genes related to anthocyanin biosynthesis had higher expression level in purple than white except F3'H, which implied that anthocyanin biosynthesis was related to purple grain traits of tetraploid wheat. The MYC transcription factor Unigene25029_All was located on chromosome 2 AL, and its expression level in purple pericarp was 4.78 times higher than that in white pericarp, and homologous to TaMYC1(pp3), which controls purple pericarp in common wheat. This gene was likely to be a candidate for purple pericarp traits in teraploid wheat. In this article, candidate gene for purple pericarp traits in teraploid wheat was screened, which might be helpful to understand the origin and molecular mechanism of purple pericarp traits in wheat.
引文
Arbuzova V.S.,Maystrenko O.I.,and Popova O.M..,1998,Development of near-isogenic lines of the common wheat cultivar'Saratovskaya 29',Cereal Res.Commun.,26(1):39-46
Dobrovolskaya O.,Arbuzova V.S.,Lohwasser U.,Roeder M.S.,and Borner A.,2006,Microsatellite mapping of complementary genes for purple grain colour in bread wheat(Triticum aestivum L.),Euphytica,150(3):355-364
Dorn K.M.,Fankhauser J.D.,Wyse D.L.,and Marks M.D.,2013,De novo assembly of the pennycress(Thlaspi arvense)transcriptome provides tools for the development of a winter cover crop and biodiesel feedstock,Plant J.,75(6):1028-1038
Elsm A.A.,Abouarab A.A.,Gamel T.H.,Hucl P.,Young J.C.,and Rabalski I.,2008,Fractionation of bl ue wheat anthocyanin compounds and their contribution to antioxidant properties,J.Agric.Food Chem.,56(23):11171-11177
Elsm A.A.,Young J.C.,and Rabalski I.,2006,Anthocyanin composition in black,blue,pink,purple,and red cereal grains,J.Agric.Food Chem.,54(13):4696-4704
Firon N.,LaBonte D.,Villordon A.,Kfir Y.,Solis J.,Lapis E.,Perlman T.S.,Doron-Faigenboim A.,Hetzroni A.,Althan L.,and Nadir L.A.,2013,Transcriptional profiling of sweetpotato(Ipomoea batatas)roots indicates down-regulation of lignin biosynthesis and up-regulation of starch biosynthesis at an early stage of storage root formation,BMC Genomics,14:460
Gion K.,Suzuri R.,Ishiguro K.,Katsumoto Y.,Tsuda S.,Tanaka Y.,Mouradova E.,Brugliera F.,and Chandler S.,2012,Genetic engineering of floricultural crops:modification of flower colour,flowering and shape,Acta Horticulturae,953(953):209-216
Grabherr M.G.,Haas B.J.,Yassour M.,Levin J.Z.,Thompson D.A.,Amit I.,Adiconis X.,Fan L.,Raychowdhury R.,Zeng Q.D.,Chen Z.H.,Mauce li E.,Hacohen N.,Gnirke A.,Rhind N.,Palma F.,Birren B.W.,Nusbaum C.,Lindblad-Toh K.,Friedman N.,and Regev A.,2011,Trinity:reconstructing a full-length transcriptome without a genome from RNA-Seq data,Nat.Biotechnol.,29(7):644
Guo J.,Han W.,and Wang M.H.,2008,Ultraviolet and environmental stresses involved in the induction and regulation of anthocyanin bio synthesis:a review,Afr.J.Biotechnol.,7(25):4966-4972
Hu J.P.,Anderson B.,and Wessler S.R.,1996,Isolation and characterization of rice R genes:Evidence for distinct evolutionary paths in rice and maize,Genetics,142(3):1021-1031
Iseli C.,Jongeneel C.V.,and Bucher P.,1999,ESTScan:a program for detecting,evaluating,and reconstructing potential coding regions in EST sequences,Intelligent Systems for Molecular Biology,99(2):138-148
Kanehisa M.,Araki M.,Goto S.,Hattori M.,Hirakawa M.,Itoh M.,Katayama T.,Kawashima S.,Okuda S.,Tokimatsu T.,and Yamanishi Y.,2008,KEGG for linking genomes to life and the environment,Nucleic Acids Res.,36(D):480-484
Khlestkina E.K.,2013,Genes determining the coloration of different organs in wheat,Russ.J.Genet.Appl.Res.,3(1):54-65
Khlestkina E.K.,Pestsova E.G.,Roder M.S.,and Borner A.,2002,Molecular mapping,phenotypic expression and geographical distribution of genes determining anthocyanin pigmentation of coleoptiles in wheat(Triticum aestivum L.),Theor.Appl.Genet.,104(4):632-637
Khlestkina E.K.,Roder M.S.,and Borner A.,2010,Mapping genes controlling anthocyanin pigmentation on the glume and pericarp in tetraploid wheat(Triticum durum L.),Euphytica,171(1):65-69
Li N.,Li S.M.,Zhang K.P.,Chen W.J.,Zhang B.,Wang D.W.,Liu D.C.,Liu B.L.,and Zhang H.G.,2017,ThMYC4E,candidate Blue aleurone 1 gene controlling the associated trait in Triticum aestivum,PLoS One,12(7):e0181116
Ludwig S.R.,Habera L.F.,Dellaporta S.L.,and Wessler S.R.,1989,Lc,a member of the maize R gene family responsible for tissue-sprcific anthocyanin production,encodes a protein simila to transcriptional activators and contains the myc-homology region,Proc.Natl.Acad.Sci.USA,86(18):7092-7096
Neuman P.R.,Waines J.G.,Hilu K.W.,and Barnhart D.,1983,Chromosomal location of genes-controlling flavonoid production in hexaploid wheat,Genetics,103(2):313-321
Romualdi C.,Bortoluzzi S.,D'alessi F.,and Danieli G.A.,2003,IDEG6:a web tool for detection of differentially expressed genes in multiple tag sampling experiments,Physiol.Genomics,12(2):159-162
Saito K.,Yonekura-Sakakibara K.,Nakabayashi R.,Higashi Y.,Yamazaki M.,Tohge T.,and Fernie A.R.,2013,The flavonoid biosynthetic pathway in Arabidopsis:Structural and genetic diversity,Plant Physiol.Biochem.,72:21-34
Saitoh K.,Onishi K.,Mikami I.,Thidar K.,and Sano Y.,2004,Allelic diversification at the C(OsC1)locus of wild and cultivated rice:Nucleotide changes associated with phenotypes,Genetics,168(2):997-1007
Sharman B.C.,1958,'Purple pericarp':a monofactorial dominant in tetraploid wheats,Nature,181(181):929
Shoeva O.Y.,Gordeeva E.I.,and Khlestkina E.K.,2014,The regulation of anthocyanin synthesis in the wheat pericarp,Molecules,19(12):20266-20279
Tereshchenko O.Y.,Arbuzova V.S.,and Khlestkina E.K.,2013,Allelic state of the genes conferring purple pigmentation in different wheat organs predetermines transcriptional activity of the anthocyanin bi osynthesis structural genes,J.Cereal Sci.,57(1):10-13
Tereshchenko O.Y.,Gordeeva E.I.,Arbuzova V.S.,Borner A.,and Khlestkina E.K.,2012,The D genome carries a gene determining purple grain colour in wheat,Cereal Res.Commun.,40(3):334-341
Wang C.X.,and Shu Q.Y.,2007,Fine mapping and candidate gene analysis of purple pericarp gene Pb in rice(Oryza sativa L.),Chinese Science Bulletin,52(22):3097-3104
Xu W.J.,Dubos C.,and Lepiniec L.,2015,Transcriptional control of flavonoid biosynthesis by MYB-bHLH-WDR complexes,Trends Plant Sci.,20(3):176-185
Zeven A.C.,1991,Wheats with purple and blue grains:a review,Euphytica,56(3):243-258
Zhang D.J.,Yuan W.T.,Li M.T.,Liu M.C.,and Zhang Y.H.,2015,Correlation analysis between gene polymorphism of trnS-G sequence and content of salidrosides in Tibetan medicine of Rhodiola taohoensis S.H.Fu,Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),34(1):130-135(张得钧,元王涛,李梦婷,刘明成,张育浩,2015,藏药洮河红景天trnS-G基因与红景天苷含量变异的相关性研究,基因组学与应用生物学,34(1):130-135)
Zhang N.,Liu B.L.,Ma C.Y.,Zhang G.D.,Chang J.,Si H.J.,and Wang D.,2014a,Transcriptome characterization and sequencing-based identification of drought-responsive genes in potato,Mol.Biol.Rep.,41(1):505-517
Zhang Y.,Butelli E.,and Martin C.,2014b,Engineering anthocyanin biosynthesis in plants,Curr.Opin.Plant Biol.,19:81-90
Zong Y.,Xi X.Y.,Li S.M.,Chen W.J.,Zhang B.,Liu D.C.,Liu B.L.,Wang D.W.,and Zhang H.G.,2017,Allelic variation and transcriptional isoforms of wheat TaMYC1 gene regulating anthocyanin synthesis in pericarp,Front.Plant Sci.,8:1645
Dobrovolskaya O.,Arbuzova V.S.,Lohwasser U.,Roeder M.S.,and Borner A.,2006,Microsatellite mapping of complementary genes for purple grain colour in bread wheat(Triticum aestivum L.),Euphytica,150(3):355-364
Dorn K.M.,Fankhauser J.D.,Wyse D.L.,and Marks M.D.,2013,De novo assembly of the pennycress(Thlaspi arvense)transcriptome provides tools for the development of a winter cover crop and biodiesel feedstock,Plant J.,75(6):1028-1038
Elsm A.A.,Abouarab A.A.,Gamel T.H.,Hucl P.,Young J.C.,and Rabalski I.,2008,Fractionation of bl ue wheat anthocyanin compounds and their contribution to antioxidant properties,J.Agric.Food Chem.,56(23):11171-11177
Elsm A.A.,Young J.C.,and Rabalski I.,2006,Anthocyanin composition in black,blue,pink,purple,and red cereal grains,J.Agric.Food Chem.,54(13):4696-4704
Firon N.,LaBonte D.,Villordon A.,Kfir Y.,Solis J.,Lapis E.,Perlman T.S.,Doron-Faigenboim A.,Hetzroni A.,Althan L.,and Nadir L.A.,2013,Transcriptional profiling of sweetpotato(Ipomoea batatas)roots indicates down-regulation of lignin biosynthesis and up-regulation of starch biosynthesis at an early stage of storage root formation,BMC Genomics,14:460
Gion K.,Suzuri R.,Ishiguro K.,Katsumoto Y.,Tsuda S.,Tanaka Y.,Mouradova E.,Brugliera F.,and Chandler S.,2012,Genetic engineering of floricultural crops:modification of flower colour,flowering and shape,Acta Horticulturae,953(953):209-216
Grabherr M.G.,Haas B.J.,Yassour M.,Levin J.Z.,Thompson D.A.,Amit I.,Adiconis X.,Fan L.,Raychowdhury R.,Zeng Q.D.,Chen Z.H.,Mauce li E.,Hacohen N.,Gnirke A.,Rhind N.,Palma F.,Birren B.W.,Nusbaum C.,Lindblad-Toh K.,Friedman N.,and Regev A.,2011,Trinity:reconstructing a full-length transcriptome without a genome from RNA-Seq data,Nat.Biotechnol.,29(7):644
Guo J.,Han W.,and Wang M.H.,2008,Ultraviolet and environmental stresses involved in the induction and regulation of anthocyanin bio synthesis:a review,Afr.J.Biotechnol.,7(25):4966-4972
Hu J.P.,Anderson B.,and Wessler S.R.,1996,Isolation and characterization of rice R genes:Evidence for distinct evolutionary paths in rice and maize,Genetics,142(3):1021-1031
Iseli C.,Jongeneel C.V.,and Bucher P.,1999,ESTScan:a program for detecting,evaluating,and reconstructing potential coding regions in EST sequences,Intelligent Systems for Molecular Biology,99(2):138-148
Kanehisa M.,Araki M.,Goto S.,Hattori M.,Hirakawa M.,Itoh M.,Katayama T.,Kawashima S.,Okuda S.,Tokimatsu T.,and Yamanishi Y.,2008,KEGG for linking genomes to life and the environment,Nucleic Acids Res.,36(D):480-484
Khlestkina E.K.,2013,Genes determining the coloration of different organs in wheat,Russ.J.Genet.Appl.Res.,3(1):54-65
Khlestkina E.K.,Pestsova E.G.,Roder M.S.,and Borner A.,2002,Molecular mapping,phenotypic expression and geographical distribution of genes determining anthocyanin pigmentation of coleoptiles in wheat(Triticum aestivum L.),Theor.Appl.Genet.,104(4):632-637
Khlestkina E.K.,Roder M.S.,and Borner A.,2010,Mapping genes controlling anthocyanin pigmentation on the glume and pericarp in tetraploid wheat(Triticum durum L.),Euphytica,171(1):65-69
Li N.,Li S.M.,Zhang K.P.,Chen W.J.,Zhang B.,Wang D.W.,Liu D.C.,Liu B.L.,and Zhang H.G.,2017,ThMYC4E,candidate Blue aleurone 1 gene controlling the associated trait in Triticum aestivum,PLoS One,12(7):e0181116
Ludwig S.R.,Habera L.F.,Dellaporta S.L.,and Wessler S.R.,1989,Lc,a member of the maize R gene family responsible for tissue-sprcific anthocyanin production,encodes a protein simila to transcriptional activators and contains the myc-homology region,Proc.Natl.Acad.Sci.USA,86(18):7092-7096
Neuman P.R.,Waines J.G.,Hilu K.W.,and Barnhart D.,1983,Chromosomal location of genes-controlling flavonoid production in hexaploid wheat,Genetics,103(2):313-321
Romualdi C.,Bortoluzzi S.,D'alessi F.,and Danieli G.A.,2003,IDEG6:a web tool for detection of differentially expressed genes in multiple tag sampling experiments,Physiol.Genomics,12(2):159-162
Saito K.,Yonekura-Sakakibara K.,Nakabayashi R.,Higashi Y.,Yamazaki M.,Tohge T.,and Fernie A.R.,2013,The flavonoid biosynthetic pathway in Arabidopsis:Structural and genetic diversity,Plant Physiol.Biochem.,72:21-34
Saitoh K.,Onishi K.,Mikami I.,Thidar K.,and Sano Y.,2004,Allelic diversification at the C(OsC1)locus of wild and cultivated rice:Nucleotide changes associated with phenotypes,Genetics,168(2):997-1007
Sharman B.C.,1958,'Purple pericarp':a monofactorial dominant in tetraploid wheats,Nature,181(181):929
Shoeva O.Y.,Gordeeva E.I.,and Khlestkina E.K.,2014,The regulation of anthocyanin synthesis in the wheat pericarp,Molecules,19(12):20266-20279
Tereshchenko O.Y.,Arbuzova V.S.,and Khlestkina E.K.,2013,Allelic state of the genes conferring purple pigmentation in different wheat organs predetermines transcriptional activity of the anthocyanin bi osynthesis structural genes,J.Cereal Sci.,57(1):10-13
Tereshchenko O.Y.,Gordeeva E.I.,Arbuzova V.S.,Borner A.,and Khlestkina E.K.,2012,The D genome carries a gene determining purple grain colour in wheat,Cereal Res.Commun.,40(3):334-341
Wang C.X.,and Shu Q.Y.,2007,Fine mapping and candidate gene analysis of purple pericarp gene Pb in rice(Oryza sativa L.),Chinese Science Bulletin,52(22):3097-3104
Xu W.J.,Dubos C.,and Lepiniec L.,2015,Transcriptional control of flavonoid biosynthesis by MYB-bHLH-WDR complexes,Trends Plant Sci.,20(3):176-185
Zeven A.C.,1991,Wheats with purple and blue grains:a review,Euphytica,56(3):243-258
Zhang D.J.,Yuan W.T.,Li M.T.,Liu M.C.,and Zhang Y.H.,2015,Correlation analysis between gene polymorphism of trnS-G sequence and content of salidrosides in Tibetan medicine of Rhodiola taohoensis S.H.Fu,Jiyinzuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology),34(1):130-135(张得钧,元王涛,李梦婷,刘明成,张育浩,2015,藏药洮河红景天trnS-G基因与红景天苷含量变异的相关性研究,基因组学与应用生物学,34(1):130-135)
Zhang N.,Liu B.L.,Ma C.Y.,Zhang G.D.,Chang J.,Si H.J.,and Wang D.,2014a,Transcriptome characterization and sequencing-based identification of drought-responsive genes in potato,Mol.Biol.Rep.,41(1):505-517
Zhang Y.,Butelli E.,and Martin C.,2014b,Engineering anthocyanin biosynthesis in plants,Curr.Opin.Plant Biol.,19:81-90
Zong Y.,Xi X.Y.,Li S.M.,Chen W.J.,Zhang B.,Liu D.C.,Liu B.L.,Wang D.W.,and Zhang H.G.,2017,Allelic variation and transcriptional isoforms of wheat TaMYC1 gene regulating anthocyanin synthesis in pericarp,Front.Plant Sci.,8:1645