紫背天葵及其近缘种白子菜花青素合成相关基因分析
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摘要
【目的】获取紫背天葵花青素合成代谢核心酶系列信息,为进一步从事紫背天葵或其他特色植物中花青素合成代谢相关基础研究提供依据。【方法】以紫背天葵为材料,以其近缘种白子菜为对照,进行转录组测序分析。【结果】在获取的80 798条单基因簇中,通过8个数据库比对分析,共有43 369条注释到相关信息。这些单基因簇在NR及Swiss-Prot这2个数据库中注释到花青素合成代谢相关系列酶为104个,具体包括10个查尔酮合成酶、5个查尔酮异构酶、36个黄烷酮3-羟化酶、19个二氢黄酮醇4-还原酶、9个花青素合成酶和25个类黄酮3-O-糖基转移酶。通过FPKM均值统计分析可知,这6大家族酶在该两种野菜中表达量差异明显的分别有1、1、3、4、2、4个。在这15个差异表达明显基因中,上调基因7个、下调基因8个。【结论】获取的紫背天葵及其近缘种白子菜中的104个相关酶系列信息中,有15个与紫背天葵花青素合成代谢密切相关。
【Objective】The core enzymes in anthocyanin anabolism were investigated,to provid a reference for further studies on the anthocyanin metabolism of G.bicolorand similar plants.【Method】Transcriptome of Gynura bicolor and G.divaricata were sequenced using the Illumina HiSeq 2 500 platform for the study.【Result】The Denovo mapping on 8 databases showed that 43 369 out of 80 798 unigenes annotated differences between the two wild vegetables.Among them,104 unigenes were identified to relate to the anthocyanin anabolism based on two databases on the non-redundant and Swiss-Prot proteins.They included 10 chalcone synthases,5 chalcone isomerases,36 flavanone 3-hydroxylases,19 dihydroflavonols,9 anthocyanidin synthases,and 25 flavonoid 3-Oglucosyl transferases.According to the average FPKMs,7 of the 15 differentially expressed genes(i.e.,1,1,3,4,2,and 4 genes from each of the 6 enzyme categories)between the 2 wild vegetables were found to be upregulated,and 8 down-regulated.【Conclusion】There were 15 enzymes in the 104 unigenes appeared to closely associate with the anthocyanin synthesis in G.bicolor and G.divaricata.
【Objective】The core enzymes in anthocyanin anabolism were investigated,to provid a reference for further studies on the anthocyanin metabolism of G.bicolorand similar plants.【Method】Transcriptome of Gynura bicolor and G.divaricata were sequenced using the Illumina HiSeq 2 500 platform for the study.【Result】The Denovo mapping on 8 databases showed that 43 369 out of 80 798 unigenes annotated differences between the two wild vegetables.Among them,104 unigenes were identified to relate to the anthocyanin anabolism based on two databases on the non-redundant and Swiss-Prot proteins.They included 10 chalcone synthases,5 chalcone isomerases,36 flavanone 3-hydroxylases,19 dihydroflavonols,9 anthocyanidin synthases,and 25 flavonoid 3-Oglucosyl transferases.According to the average FPKMs,7 of the 15 differentially expressed genes(i.e.,1,1,3,4,2,and 4 genes from each of the 6 enzyme categories)between the 2 wild vegetables were found to be upregulated,and 8 down-regulated.【Conclusion】There were 15 enzymes in the 104 unigenes appeared to closely associate with the anthocyanin synthesis in G.bicolor and G.divaricata.
引文
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[6]贾赵东,马佩勇,边小峰,等.植物花青素合成代谢途径及其分子调控[J].西北植物学报,2014,34(7):1496-1506.JIA Z D,MA P Y,BIAN X F,et al.Biosynthesis metabolic pathway and molecular regulation of plants anthocyanin[J].Acta Botanica Boreali-Occidentalia Sinica,2014,34(7):1496-1506.(in Chinese)
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[9]CARLETTI G,LUCIN L,BUSCONI M,et al.Insight into the role of anthocyanin biosynthesis-related genes in Medicago truncatula mutants impaired in pigmentation in leaves[J].Plant Physiol Biochem,2013,70(9):123-132.
[10]KALT W,MCDONALD J E,LIU Y,et al.Flavonoid metabolites in human urine during blueberry anthocyanin intake[J].J Agric Food Chem,2017,65(8):1582-1591.
[11]戴思兰,洪艳.基于花青素苷合成和呈色机理的观赏植物花色改良分子育种[J].中国农业科学,2016,49(3):529-542.DAI S L,HONG Y.Molecular breeding for flower colors modification on ornamental plants based on the mechanism of anthocyanins biosynthesis and coloration[J].Scientia Agricultura Sinica,2016,49(3):529-542.(in Chinese)
[12]李卫星,杨舜博,何智冲,等.植物叶色变化机制研究进展[J].园艺学报,2017,44(9):1811-1824.LI W X,YANG S B,HE Z C,et al.Research advances in the regulatory mechanisms of leaf coloration[J].Acta Horticulturae Sinica,2017,44(9):1811-1824.(in Chinese)
[13]孙玉燕,段蒙蒙,邱杨,等.心里美萝卜花青素合成酶基因RsANS克隆及花青素生物合成相关基因表达分析[J].植物遗传资源学报,2016,17(5):889-896.SUN Y Y,DUAN M M,QIU Y,et al.Anthocyanidin Synthase Gene Cloning and Expression Analysis of Anthocyanidin Biosynthesis Related Genes in'Xinlimei'Radish(Raphanus sativus L.)[J].Journal of Genetic Resources,2016,17(5):889-896.(in Chinese)
[14]SPRINGOB K,NAKAJIMA J,YAMAZAKIi M,et al.Recent advances in the biosynthesis and accumulation of anthocyanins[J].Natural Product Reports,2003,20(3):288-303.
[15]CHEN G,LIU H,WEI Q,et al.The acyl-activating enzyme PhAAE13is an alternative enzymatic source of precursors for anthocyanin biosynthesis in petunia flowers[J].J Exp Bot,2017,68(3):457-467.
[16]GAO C,LI D,JIN C,et al.Genome-wide identification of GLABRA3 downstream genes for anthocyanin biosynthesis and trichome formation in Arabidopsis[J].Biochem Biophys Res Commun,2017,485(2):360-365.
[17]MORITA Y,ISHIGURO K,TANAKA Y,et al.Spontaneous mutations of the UDP-glucose:flavonoid 3-O-glucosyltransferase gene confers pale-and dull-colored flowers in the Japanese and common morning glories[J].Planta,2015,242(3):575-587.
[18]LI X G,WANG J,YU Z Y.Cloning of an anthocyanidin synthase gene homolog from blackcurrant(Ribes nigrum L.)and its expression at different fruit stages[J].Genet Mol Res,2015,14(1):2726-2734.
[19]CHU Y X,CHEN H R,WU A Z,et al.Expression analysis of dihydroflavonol 4-reductase genes in Petunia hybrida[J].Genet Mol Res,2015,14(2):5010-5021.
[20]SANGWAN R S,TRIPATHI S,SINGH J,et al.De novo sequencing and assembly of Centella asiatica leaf transcriptome for mapping of structural,functional and regulatory genes with special reference to secondary metabolism[J].Gene,2013,525(2):58-76.
[21]张少平,邱珊莲,郑云云,等.紫色黄秋葵转录组功能基因测序及分析[J].核农学报,2017,31(4):643-653.ZHANG S P,QIU S L,ZHENG Y Y,et al.The Gynura bicolor Transcriptome as a Source for Anthocyanidin Gene Sequence Analysis[J].Journal of Nuclear Agricultural Sciences,2017,31(4):643-653.(in Chinese)
[22]KAYMAZ Y,ODUOR C I,YU H,et al.Comprehensive Transcriptome and Mutational Profiling of Endemic Burkitt Lymphoma Reveals EBV Type-Specific Differences[J].Mol Cancer Res,2017,15(5):563-576.
[23]张少平,邱珊莲,邓源,等.紫背天葵花青素相关研究与运用[J].中国农学通报,2015,31(22):157-162.ZHANG S P,QIU S L,DENG Y,et al.Study and application of the anthocyanin from Gynum bicolor[J].Chinese Agricultural Science Bulletin,2015,31(22):157-162.(in Chinese)
[24]张少平,徐强,张帅,等.药食同源蔬菜-白子菜[J].蔬菜,2016,308(8):74-75.ZHANG S P,XU Q,ZHANG S,et al.The Gynura divaricata as medicinal and edible vegetable[J].Journal of Vegetable,2016,308(8):74-75.(in Chinese)
[25]张少平,张帅,郑开斌,等.紫背天葵种苗组培快繁技术[J].福建农业科技,2017,48(9):32-33.ZHANG S P,ZHANG S,ZHENG K B,et al.Seedling production to the Gynura bicolor by tissue culture and rapid propagation[J].Fujian Agriculture Science and Technology,2017,48(9):32-33.(in Chinese)
[26]ALTSCHUL S F,MADDEN T L,SCHAFFER A A,et al.Gapped BLAST and PSI BLAST:a new generation of protein database search programs[J].Nucleic Acids Research Italic,1997,25(17):3389-3402.
[27]ANN K H,ALBERTO L E,JOHN P,et al.De Novo transcriptome assembly and sex-biased gene expression in the cyclical parthenogenetic daphnia galeata[J].Genome Biol Evol,2016,8(10):3120-3139.
[28]NINA F O,LAUREN A O,STEPHEN J B,et al.Optimization of next-generation sequencing transcriptome annotation for species lacking sequenced genomes[J].Mol Ecol Resour,2016,16(2):446-458.
[29]HUIAI G,QI P,YANG H,et al.Characteristics ofα-Gal epitope,anti-Gal antibody,α-1,3galactosyltransferase and its clinical exploitation[J].Int J Mol Med,2016,37(1):11-20.
[30]LIM S H,YOU M K,KIM D H,et al.RNAi-mediated suppression of dihydroflavonol 4-reductase in tobacco allows fine-tuning of flower color and flux through the flavonoid biosynthetic pathway[J].Plant Physiol Biochem,2016,109(10):482-490.
[31]MORITA Y,ISHIGURO K,TANAKA Y,et al.Spontaneous mutations of the UDP-glucose:flavonoid 3-O-glucosyltransferase gene confers pale-and dull-colored flowers in the Japanese and common morning glories[J].Planta,2015,242(3):575-587.
[32]张少平,洪建基,邱珊莲,等.紫背天葵高通量转录组测序分析[J].园艺学报,2016,43(5):935-946.ZHANG S P,HONG J J,QIU S L,et al.Sequencing and Analysis of the Transcriptome of Gynura bicolor[J].Acta Horticulturae Sinica,2016,43(5):935-946.(in Chinese)
[33]张少平,张少华,邱珊莲,等.基于转录组测序的紫背天葵花青素相关基因分析[J].核农学报,2018,32(4):639-645.ZHANG S P,ZHANG S H,QIU S L,et al.The Gynura bicolor transcriptome as A source for anthocyanidin gene sequence analysis[J].Journal of Nuclear Agricultural Sciences,2018,32(4):639-645.(in Chinese)
[2]CHEN T,HU S,ZHANG H,et al.Anti-inflammatory effects of Dioscorea alata L.anthocyanins in a TNBS-induced colitis model[J].Food Funct,2017,8(2):659-669.
[3]GULDIKEN B,GIBIS M,BOYACIOGLU D,et al.Impact of liposomal encapsulation on degradation of anthocyanins of black carrot extract by adding ascorbic acid[J].Food Funct,2017,8(3):1085-1093.
[4]SENEM K,ESRA C,CHARLOTTE G,et al.Anthocyanin absorption and metabolism by human intestinal caco-2cells-a review[J].Int J Mol Sci,2015,16(9):21555-21574.
[5]VENANCIO V P,CIPRIANO P A,KIM H,et al.Cocoplum(Chryso balanus icaco L.)anthocyanins exert antiinflammatory activity in human colon cancer and non-malignant colon cells[J].Food Funct,2017,8(1):307-314.
[6]贾赵东,马佩勇,边小峰,等.植物花青素合成代谢途径及其分子调控[J].西北植物学报,2014,34(7):1496-1506.JIA Z D,MA P Y,BIAN X F,et al.Biosynthesis metabolic pathway and molecular regulation of plants anthocyanin[J].Acta Botanica Boreali-Occidentalia Sinica,2014,34(7):1496-1506.(in Chinese)
[7]KATARINA J,JANKA B,MICHAELA H,et al.The effects of anthocyanin-rich wheat diet on the oxidative status and behavior of rats[J].Croat Med J,2016,57(2):119-129.
[8]赵启明,李范,李萍.花青素生物合成关键酶的研究进展[J].生物技术通报,2012(12):25-32.ZHAO Q M,LI F,LI P.Research advances on core enzymes of anthocyanidin biosynthesis[J].Biotechnology Bulletin,2012(12):25-32.(in Chinese)
[9]CARLETTI G,LUCIN L,BUSCONI M,et al.Insight into the role of anthocyanin biosynthesis-related genes in Medicago truncatula mutants impaired in pigmentation in leaves[J].Plant Physiol Biochem,2013,70(9):123-132.
[10]KALT W,MCDONALD J E,LIU Y,et al.Flavonoid metabolites in human urine during blueberry anthocyanin intake[J].J Agric Food Chem,2017,65(8):1582-1591.
[11]戴思兰,洪艳.基于花青素苷合成和呈色机理的观赏植物花色改良分子育种[J].中国农业科学,2016,49(3):529-542.DAI S L,HONG Y.Molecular breeding for flower colors modification on ornamental plants based on the mechanism of anthocyanins biosynthesis and coloration[J].Scientia Agricultura Sinica,2016,49(3):529-542.(in Chinese)
[12]李卫星,杨舜博,何智冲,等.植物叶色变化机制研究进展[J].园艺学报,2017,44(9):1811-1824.LI W X,YANG S B,HE Z C,et al.Research advances in the regulatory mechanisms of leaf coloration[J].Acta Horticulturae Sinica,2017,44(9):1811-1824.(in Chinese)
[13]孙玉燕,段蒙蒙,邱杨,等.心里美萝卜花青素合成酶基因RsANS克隆及花青素生物合成相关基因表达分析[J].植物遗传资源学报,2016,17(5):889-896.SUN Y Y,DUAN M M,QIU Y,et al.Anthocyanidin Synthase Gene Cloning and Expression Analysis of Anthocyanidin Biosynthesis Related Genes in'Xinlimei'Radish(Raphanus sativus L.)[J].Journal of Genetic Resources,2016,17(5):889-896.(in Chinese)
[14]SPRINGOB K,NAKAJIMA J,YAMAZAKIi M,et al.Recent advances in the biosynthesis and accumulation of anthocyanins[J].Natural Product Reports,2003,20(3):288-303.
[15]CHEN G,LIU H,WEI Q,et al.The acyl-activating enzyme PhAAE13is an alternative enzymatic source of precursors for anthocyanin biosynthesis in petunia flowers[J].J Exp Bot,2017,68(3):457-467.
[16]GAO C,LI D,JIN C,et al.Genome-wide identification of GLABRA3 downstream genes for anthocyanin biosynthesis and trichome formation in Arabidopsis[J].Biochem Biophys Res Commun,2017,485(2):360-365.
[17]MORITA Y,ISHIGURO K,TANAKA Y,et al.Spontaneous mutations of the UDP-glucose:flavonoid 3-O-glucosyltransferase gene confers pale-and dull-colored flowers in the Japanese and common morning glories[J].Planta,2015,242(3):575-587.
[18]LI X G,WANG J,YU Z Y.Cloning of an anthocyanidin synthase gene homolog from blackcurrant(Ribes nigrum L.)and its expression at different fruit stages[J].Genet Mol Res,2015,14(1):2726-2734.
[19]CHU Y X,CHEN H R,WU A Z,et al.Expression analysis of dihydroflavonol 4-reductase genes in Petunia hybrida[J].Genet Mol Res,2015,14(2):5010-5021.
[20]SANGWAN R S,TRIPATHI S,SINGH J,et al.De novo sequencing and assembly of Centella asiatica leaf transcriptome for mapping of structural,functional and regulatory genes with special reference to secondary metabolism[J].Gene,2013,525(2):58-76.
[21]张少平,邱珊莲,郑云云,等.紫色黄秋葵转录组功能基因测序及分析[J].核农学报,2017,31(4):643-653.ZHANG S P,QIU S L,ZHENG Y Y,et al.The Gynura bicolor Transcriptome as a Source for Anthocyanidin Gene Sequence Analysis[J].Journal of Nuclear Agricultural Sciences,2017,31(4):643-653.(in Chinese)
[22]KAYMAZ Y,ODUOR C I,YU H,et al.Comprehensive Transcriptome and Mutational Profiling of Endemic Burkitt Lymphoma Reveals EBV Type-Specific Differences[J].Mol Cancer Res,2017,15(5):563-576.
[23]张少平,邱珊莲,邓源,等.紫背天葵花青素相关研究与运用[J].中国农学通报,2015,31(22):157-162.ZHANG S P,QIU S L,DENG Y,et al.Study and application of the anthocyanin from Gynum bicolor[J].Chinese Agricultural Science Bulletin,2015,31(22):157-162.(in Chinese)
[24]张少平,徐强,张帅,等.药食同源蔬菜-白子菜[J].蔬菜,2016,308(8):74-75.ZHANG S P,XU Q,ZHANG S,et al.The Gynura divaricata as medicinal and edible vegetable[J].Journal of Vegetable,2016,308(8):74-75.(in Chinese)
[25]张少平,张帅,郑开斌,等.紫背天葵种苗组培快繁技术[J].福建农业科技,2017,48(9):32-33.ZHANG S P,ZHANG S,ZHENG K B,et al.Seedling production to the Gynura bicolor by tissue culture and rapid propagation[J].Fujian Agriculture Science and Technology,2017,48(9):32-33.(in Chinese)
[26]ALTSCHUL S F,MADDEN T L,SCHAFFER A A,et al.Gapped BLAST and PSI BLAST:a new generation of protein database search programs[J].Nucleic Acids Research Italic,1997,25(17):3389-3402.
[27]ANN K H,ALBERTO L E,JOHN P,et al.De Novo transcriptome assembly and sex-biased gene expression in the cyclical parthenogenetic daphnia galeata[J].Genome Biol Evol,2016,8(10):3120-3139.
[28]NINA F O,LAUREN A O,STEPHEN J B,et al.Optimization of next-generation sequencing transcriptome annotation for species lacking sequenced genomes[J].Mol Ecol Resour,2016,16(2):446-458.
[29]HUIAI G,QI P,YANG H,et al.Characteristics ofα-Gal epitope,anti-Gal antibody,α-1,3galactosyltransferase and its clinical exploitation[J].Int J Mol Med,2016,37(1):11-20.
[30]LIM S H,YOU M K,KIM D H,et al.RNAi-mediated suppression of dihydroflavonol 4-reductase in tobacco allows fine-tuning of flower color and flux through the flavonoid biosynthetic pathway[J].Plant Physiol Biochem,2016,109(10):482-490.
[31]MORITA Y,ISHIGURO K,TANAKA Y,et al.Spontaneous mutations of the UDP-glucose:flavonoid 3-O-glucosyltransferase gene confers pale-and dull-colored flowers in the Japanese and common morning glories[J].Planta,2015,242(3):575-587.
[32]张少平,洪建基,邱珊莲,等.紫背天葵高通量转录组测序分析[J].园艺学报,2016,43(5):935-946.ZHANG S P,HONG J J,QIU S L,et al.Sequencing and Analysis of the Transcriptome of Gynura bicolor[J].Acta Horticulturae Sinica,2016,43(5):935-946.(in Chinese)
[33]张少平,张少华,邱珊莲,等.基于转录组测序的紫背天葵花青素相关基因分析[J].核农学报,2018,32(4):639-645.ZHANG S P,ZHANG S H,QIU S L,et al.The Gynura bicolor transcriptome as A source for anthocyanidin gene sequence analysis[J].Journal of Nuclear Agricultural Sciences,2018,32(4):639-645.(in Chinese)