胡萝卜根中主要类胡萝卜素含量相关QTL的精细定位
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摘要
以白色的野生胡萝卜‘松滋野生’(Ws)和橘色的栽培胡萝卜品种‘Amsterdam’(Af)为亲本构建的回交重组自交系(BIL)为试材,基于低倍重测序技术开发SNP标记,构建了由1 976个Bin标记组成,包含29 435个SNP标记的遗传图谱。图谱总距离834.28 c M,平均图距0.42 c M。通过对胡萝卜肉质根中类胡萝卜素含量相关QTL分析,在连锁群LG04和LG08中检测到调控α–胡萝卜素、β–胡萝卜素、ζ–胡萝卜素、叶黄素、玉米黄质和总类胡萝卜素含量的主效QTL(M-QTL)2、2、3、2、2和2个,表型贡献率为11.47%~19.18%;另检测到调控α–胡萝卜素、β–胡萝卜素、ζ–胡萝卜素、玉米黄质和总类胡萝卜素含量的上位性QTL(E-QTL)1、1、2、1和1个,表型贡献率为2.50%~3.66%。在M-QTL显著区间内共检索到36个有功能注释的预测基因,其中Dck018297为ζ–胡萝卜素脱氢酶2基因,与调控β–胡萝卜素合成和总类胡萝卜素含量有关;Dck008006为乙烯响应因子2.2的同源基因,与调控α–胡萝卜素、ζ–胡萝卜素合成有关;Dck029898为转录因子b HLH135的同源基因,与调控玉米黄质合成有关。
Based on whole-genome resequencing data,a high resolution genetic map was developed by using a set of backcross inbred lines(BILs)crossed by a wild species‘Songzi'with a cultivar‘Amsterdam',and used to map QTLs for main carotenoids in carrot root. Finally,the genetic map was consisted of 1 976 Bins which contained 29 435 SNPs. The total genetic distance of map was 834.28 c M with an average interval of 0.42 c M between Bin markers. There were 2,2,3,2,2 and 2 main-effect QTLs(M-QTLs)associated with α-carotene,β-carotene,ζ-carotene,lutein,zeaxanthin and total carotenoid contents,respectively,with 11.47%–19.18% phenotypic variance. There were 1,1,2,1 and 1 epistatic QTLs contributed to α-carotene,β-carotene,ζ-carotene,zeaxanthin and total carotenoid contents,respectively,with 2.50%–3.66% phenotypic variance. A total of 36 predicted genes with function annotations were detected in M-QTLs. Among these,gene Dck018297 was ζ-carotene desaturase and associated with β-carotene biosynthesis and total carotenoid contents;gene Dck008006 homologouse to ERF2.2 was associated with α-carotene and zeaxanthin biosynthesis;gene Dck029898 homologouse to b HLH135 was associated with ζ-carotene biosynthesis. These three candidate genes might play central roles in the carotenoid biosynthesis in carrot root.
Based on whole-genome resequencing data,a high resolution genetic map was developed by using a set of backcross inbred lines(BILs)crossed by a wild species‘Songzi'with a cultivar‘Amsterdam',and used to map QTLs for main carotenoids in carrot root. Finally,the genetic map was consisted of 1 976 Bins which contained 29 435 SNPs. The total genetic distance of map was 834.28 c M with an average interval of 0.42 c M between Bin markers. There were 2,2,3,2,2 and 2 main-effect QTLs(M-QTLs)associated with α-carotene,β-carotene,ζ-carotene,lutein,zeaxanthin and total carotenoid contents,respectively,with 11.47%–19.18% phenotypic variance. There were 1,1,2,1 and 1 epistatic QTLs contributed to α-carotene,β-carotene,ζ-carotene,zeaxanthin and total carotenoid contents,respectively,with 2.50%–3.66% phenotypic variance. A total of 36 predicted genes with function annotations were detected in M-QTLs. Among these,gene Dck018297 was ζ-carotene desaturase and associated with β-carotene biosynthesis and total carotenoid contents;gene Dck008006 homologouse to ERF2.2 was associated with α-carotene and zeaxanthin biosynthesis;gene Dck029898 homologouse to b HLH135 was associated with ζ-carotene biosynthesis. These three candidate genes might play central roles in the carotenoid biosynthesis in carrot root.
引文
Arango J,Jourdan M,Geoffriau E,Beyer P,Welsch R.2014.Carotene hydroxylase activity determines the levels of bothα-carotene and total carotenoids in orange carrots.Plant Cell,doi:10.1105/tpc.113.122127.
Boiteux L S,Belter J G,Roberts P A,Simon P W.2000.RAPD linkage map of the genomic region encompassing the rootknot nematode(Meloidogyne javanica)resistance locus in carrot.Theor Appl Genet,100:439-446.
Briard M,Le Clerc V,Grzebelus D,Senalik D,Simon P W.2000.Modified protocols for rapid carrot genomic DNA extraction and AFLPTM analysis using silver stain or radioisotopes.Plant Mol Biol Rep,18:235-241.
Buishand J G,Gabelman W H.1979.Investigations on the inheritance of color and carotenoid content in phloem and xylem of carrot roots(Daucus carota L.).Euphytica,28:611-632.
Cavagnaro P F,Chung S M,Manin S,Yildiz M,Ali A,Alessandro M S,Iorizzo M,Senalik D A,Simon P W.2011.Microsatellite isolation and marker development in carrot-genomic distribution,linkage mapping,genetic diversity analysis and marker transferability across Apiaceae.BMC Genomics,12:386.
Cavagnaro P F,Iorizzo M,Yildiz M,Senalik D,Parsons J,Ellison S,Simon P W.2014.A gene-derived SNP-based high resolution linkage map of carrot including the location of QTL conditioning root and leaf anthocyanin pigmentation.BMC Genomics,15:1118.
Chung M Y,Vrebalov J,Alba R,Lee J,Mc Quinn R,Chung J D,Klein P,Giovannoni J.2010.A tomato(Solanum lycopersicum)APETALA2/ERFgene,Sl AP2a,is a negative regulator of fruit ripening.Plant J,64:936-947.
Endo T,Fujii H,Sugiyama A,Nakano M,Nakajima N,Ikoma Y,Omura M,Shimada T.2016.Overexpression of a citrus basic helix-loop-helix transcription factor(Cub HLH1),which is homologous to Arabidopsis activation-tagged bri1 suppressor 1 interacting factor genes,modulates carotenoid metabolism in transgenic tomato.Plant Science,243:35-48.
Goldman I L,Breitbach D N.1996.Inheritance of a recessive character controlling reduced carotenoid pigmentation in carrot(Daucus carota L.).JHered,87:380-382.
Grzebelus D,Iorizzo M,Senalik D,Ellison S,Cavagnaro P F,Macko-Podgorni A,Heller-Uszynska K,Kilian A,Nothnagel T,Allender C,Simon P W,Baranski R.2014.Diversity,genetic mapping,and signatures of domestication in the carro(tDaucus carota L.)genome,as revealed by Diversity Arrays Technology(DAr T)markers.Mol Breeding,33:625-637.
Han K,Jeong H J,Yang H B,Kang S M,Kwon J K,Kim S,Choi D,Kang B C.2016.An ultra-high-density bin map facilitates high-throughput QTL mapping of horticultural traits in pepper(Capsicum annuum).DNA Res,doi:10.1093/dnares/dsv038.
Huang X,Feng Q,Qian Q,Zhao Q,Wang L,Wang A,Guan J,Fan D,Weng Q,Huang T,Dong G,Sang T,Han B.2009.High-throughput genotyping by whole-genome resequencing.Genome Res,19:1068-1076.
Iorizzo M,Ellison S,Senalik D,Zeng P,Satapoomin P,Huang J,Bowman M,Iovene M,Sanseverino W,Cavagnaro P,Yildiz M,Spooner D M,Simon P W.2016.A high-quality carrot genome assembly provides new insights into carotenoid accumulation and asterid genome evolution.Nature Genetics,doi:10.1038/ng.3565.
Jourdan M,GagnéS,Dubois-Laurent C,Maghraoui M,Huet S,Suel A,Hamama L,Briard M,Peltier D,Geoffriau E.2015.Carotenoid content and root color of cultivated carrot:a candidate-gene association study using an original broad unstructured population.PLoS ONE,10(1):e0116674.
Just B J,Santos C A F,Fonseca M E N,Boiteux L S,Oloizia B B,Simon P W.2007.Carotenoid biosynthesis structural genes in carro(tDaucus carota):isolation,sequence-characterization,single nucleotide polymorphism(SNP)markers and genome mapping.Theor Appl Genet,114:693-704.
Lee J M,Joung J G,Mc Quinn R,Chung M Y,Fei Z,Tieman D,Klee H,Giovannoni J.2012.Combined transcriptome,genetic diversity and metabolite profiling in tomato fruit reveals that the ethylene response factor Sl ERF6 plays an important role in ripening and carotenoid accumulation.Plant J,70(2):191-204.
Li H,Durbin R.2009.Fast and accurate short read alignment with Burrows-Wheeler transform.Bioinformatics,25:1754-1760.
Liu D,Zhang J,Liu X,Wang W,Liu D,Teng Z,Fang X,Tan Z,Tang S,Yang J,Zhong J,Zhang Z.2016.Fine mapping and RNA-Seq unravels candidate genes for a major QTL controlling multiple fiber quality traits at the T1 region in upland cotton.BMC Genomics,17(1):295.
Liu T,Yu T,Xing Y.2013.Identification and validation of a yield-enhancing QTL cluster in rice(Oryza sativa L.).Euphytica,192:145-153.
Mei H W,Li Z K,Shu Q Y,Guo L B,Wang Y P,Yu X Q,Ying C S,Luo L J.2005.Gene actions of QTLs affecting several agronomic traits resolved in a recombinant inbred rice population and two backcross populations.Theor Appl Genet,110(4):649-659.
Moreno J C,Pizarro L,Fuentes P,Handford M,Cifuentes V,Stange C.2013.Levels of lycopene b-cyclase 1 modulate carotenoid gene expression and accumulation in Daucus carota.PLo S ONE,8(3):e58144.
Ou Cheng-gang,Deng Bo-tao,Bao Sheng-you,Zhao Zhi-wei,Hu Hong,Zhuang Fei-yun,Mao Shu-min.2010.QTL mapping for contents of main carotenes and lycopene in carrot(Daucus carota L.).Hereditas,32(12):1290-1295.(in Chinese)欧承刚,邓波涛,鲍生有,赵志伟,胡鸿,庄飞云,茅淑敏.2010.胡萝卜(Daucus carota L.)中主要胡萝卜素和番茄红素含量的QTL分析.遗传,32(12):1290-1295.
Qi X,Li M W,Xie M,Liu X,Ni M,Shao G,Song C,Yim K Y A,Tao Y,Wong F L,Isobe S,Wong C F,Wong K S,Xu C,Li C,Wang Y,Guan R,Sun F,Fan G,Xiao Z,Zhou F,Phang T H,Liu X,Tong S W,Chan T F,Yiu S M,Tabata S,Wang J,Xu X,Lam H M.2014.Identification of a novel salt tolerance gene in wild soybean by whole-genome sequencing.Nat Commun,5:4340.
Santos C A F,Simon P W.2002.QTL analyses reveal clustered loci for accumulation of major provitamin A carotenes and lycopene in carrot roots.Mol Genet Genomics,268:122-129.
Santos C A F,Simon P W.2006.Heritabilities and minimum gene number estimates of carrot carotenoids.Euphytica,151:79-86.
Simon P W.2000.Domestication,historical development,and modern breeding of carrot.Plant Breed Rev,19:157-190.
Simpson K,Quiroz L F,Rodriguez-Concepción M,Stange C R.2016.Differential contribution of the first two enzymes of the MEP pathway to the supply of metabolic precursors for carotenoid and chlorophyll biosynthesis in carrot(Daucus carota).Front Plant Sci,7:1344.
Umiel N,Gabelman W H.1972.Inheritance of root color and carotenoid synthesis in carrot,Daucus carota L.:orange vs.red.J Am Soc Hort Sci,97:453-460.
Van O J W.2006.JoinM ap 4.0:Software for the calculation of genetic linkage maps in experimental populations.Wageningen,Netherlands:Kyazma B.V.
Vivek B S,Simon P W.1999.Linkage relationships among molecular markers and storage root traits of carro(tDaucus carota L.ssp.sativus).Theor Appl Genet,99:58-64.
Wang H,Ou C G,Zhuang F Y,Ma Z G.2014.The dual role of phytoene synthase genes in carotenogenesis in carrot roots and leaves.Mol Breed,34:2065-2079.
Wang Hui,Ou Cheng-gang,Zhuang Fei-yun,Zhao Zhi-wei,Ma Zhen-guo.2014.Relationship of carotenoid accumulation and transcript of main genes in carotenoid biosynthesis in carrot.Acta Horticulturae Sinica,41(12):2513-2520.(in Chinese)王慧,欧承刚,庄飞云,赵志伟,马振国.2014.胡萝卜中类胡萝卜素积累与主要合成基因转录水平相关性分析.园艺学报,41(12):2513-2520.
Wang L,Wang A,Huang X,Zhao Q,Dong G,Qian Q,Sang T,Han B.2011.Mapping 49 quantitative trait loci at high resolution through sequencing-based genotyping of rice recombinant inbred lines.Theor Appl Genet,122(2):327-340.
Welsch R,Maass D,Voegel T,Dellapenna D,Beyer P.2007.Transcription factor RAP2.2 and its interacting partner SINAT2:stable elements in the carotenogenesis of Arabidopsis leaves.Plant Physiol,145(3):1073-1085.
Xu Z S,Tan H W,Wang F,Hou X L,Xiong A S.2014.Carrot DB:a genomic and transcriptomic database for carrot.Database,doi:10.1093/database/bau096.
Yang J,Hu C C,Ye X Z,Zhu J.2005.QTLNetwork 2.0.Institute of Bioinformatics,Zhejiang University,Hangzhou,China,Available:http://ibi.zju.edu.cn/software/qtlnetwork.
Zhou Z,Zhang C,Zhou Y,Hao Z,Wang Z,Zeng X,Di H,Li M,Zhang D,Yong H,Zhang S,Weng J,Li X.2016.Genetic dissection of maize plant architecture with an ultra-high density bin map based on recombinant inbred lines.BMC Genomics,17:178.
Boiteux L S,Belter J G,Roberts P A,Simon P W.2000.RAPD linkage map of the genomic region encompassing the rootknot nematode(Meloidogyne javanica)resistance locus in carrot.Theor Appl Genet,100:439-446.
Briard M,Le Clerc V,Grzebelus D,Senalik D,Simon P W.2000.Modified protocols for rapid carrot genomic DNA extraction and AFLPTM analysis using silver stain or radioisotopes.Plant Mol Biol Rep,18:235-241.
Buishand J G,Gabelman W H.1979.Investigations on the inheritance of color and carotenoid content in phloem and xylem of carrot roots(Daucus carota L.).Euphytica,28:611-632.
Cavagnaro P F,Chung S M,Manin S,Yildiz M,Ali A,Alessandro M S,Iorizzo M,Senalik D A,Simon P W.2011.Microsatellite isolation and marker development in carrot-genomic distribution,linkage mapping,genetic diversity analysis and marker transferability across Apiaceae.BMC Genomics,12:386.
Cavagnaro P F,Iorizzo M,Yildiz M,Senalik D,Parsons J,Ellison S,Simon P W.2014.A gene-derived SNP-based high resolution linkage map of carrot including the location of QTL conditioning root and leaf anthocyanin pigmentation.BMC Genomics,15:1118.
Chung M Y,Vrebalov J,Alba R,Lee J,Mc Quinn R,Chung J D,Klein P,Giovannoni J.2010.A tomato(Solanum lycopersicum)APETALA2/ERFgene,Sl AP2a,is a negative regulator of fruit ripening.Plant J,64:936-947.
Endo T,Fujii H,Sugiyama A,Nakano M,Nakajima N,Ikoma Y,Omura M,Shimada T.2016.Overexpression of a citrus basic helix-loop-helix transcription factor(Cub HLH1),which is homologous to Arabidopsis activation-tagged bri1 suppressor 1 interacting factor genes,modulates carotenoid metabolism in transgenic tomato.Plant Science,243:35-48.
Goldman I L,Breitbach D N.1996.Inheritance of a recessive character controlling reduced carotenoid pigmentation in carrot(Daucus carota L.).JHered,87:380-382.
Grzebelus D,Iorizzo M,Senalik D,Ellison S,Cavagnaro P F,Macko-Podgorni A,Heller-Uszynska K,Kilian A,Nothnagel T,Allender C,Simon P W,Baranski R.2014.Diversity,genetic mapping,and signatures of domestication in the carro(tDaucus carota L.)genome,as revealed by Diversity Arrays Technology(DAr T)markers.Mol Breeding,33:625-637.
Han K,Jeong H J,Yang H B,Kang S M,Kwon J K,Kim S,Choi D,Kang B C.2016.An ultra-high-density bin map facilitates high-throughput QTL mapping of horticultural traits in pepper(Capsicum annuum).DNA Res,doi:10.1093/dnares/dsv038.
Huang X,Feng Q,Qian Q,Zhao Q,Wang L,Wang A,Guan J,Fan D,Weng Q,Huang T,Dong G,Sang T,Han B.2009.High-throughput genotyping by whole-genome resequencing.Genome Res,19:1068-1076.
Iorizzo M,Ellison S,Senalik D,Zeng P,Satapoomin P,Huang J,Bowman M,Iovene M,Sanseverino W,Cavagnaro P,Yildiz M,Spooner D M,Simon P W.2016.A high-quality carrot genome assembly provides new insights into carotenoid accumulation and asterid genome evolution.Nature Genetics,doi:10.1038/ng.3565.
Jourdan M,GagnéS,Dubois-Laurent C,Maghraoui M,Huet S,Suel A,Hamama L,Briard M,Peltier D,Geoffriau E.2015.Carotenoid content and root color of cultivated carrot:a candidate-gene association study using an original broad unstructured population.PLoS ONE,10(1):e0116674.
Just B J,Santos C A F,Fonseca M E N,Boiteux L S,Oloizia B B,Simon P W.2007.Carotenoid biosynthesis structural genes in carro(tDaucus carota):isolation,sequence-characterization,single nucleotide polymorphism(SNP)markers and genome mapping.Theor Appl Genet,114:693-704.
Lee J M,Joung J G,Mc Quinn R,Chung M Y,Fei Z,Tieman D,Klee H,Giovannoni J.2012.Combined transcriptome,genetic diversity and metabolite profiling in tomato fruit reveals that the ethylene response factor Sl ERF6 plays an important role in ripening and carotenoid accumulation.Plant J,70(2):191-204.
Li H,Durbin R.2009.Fast and accurate short read alignment with Burrows-Wheeler transform.Bioinformatics,25:1754-1760.
Liu D,Zhang J,Liu X,Wang W,Liu D,Teng Z,Fang X,Tan Z,Tang S,Yang J,Zhong J,Zhang Z.2016.Fine mapping and RNA-Seq unravels candidate genes for a major QTL controlling multiple fiber quality traits at the T1 region in upland cotton.BMC Genomics,17(1):295.
Liu T,Yu T,Xing Y.2013.Identification and validation of a yield-enhancing QTL cluster in rice(Oryza sativa L.).Euphytica,192:145-153.
Mei H W,Li Z K,Shu Q Y,Guo L B,Wang Y P,Yu X Q,Ying C S,Luo L J.2005.Gene actions of QTLs affecting several agronomic traits resolved in a recombinant inbred rice population and two backcross populations.Theor Appl Genet,110(4):649-659.
Moreno J C,Pizarro L,Fuentes P,Handford M,Cifuentes V,Stange C.2013.Levels of lycopene b-cyclase 1 modulate carotenoid gene expression and accumulation in Daucus carota.PLo S ONE,8(3):e58144.
Ou Cheng-gang,Deng Bo-tao,Bao Sheng-you,Zhao Zhi-wei,Hu Hong,Zhuang Fei-yun,Mao Shu-min.2010.QTL mapping for contents of main carotenes and lycopene in carrot(Daucus carota L.).Hereditas,32(12):1290-1295.(in Chinese)欧承刚,邓波涛,鲍生有,赵志伟,胡鸿,庄飞云,茅淑敏.2010.胡萝卜(Daucus carota L.)中主要胡萝卜素和番茄红素含量的QTL分析.遗传,32(12):1290-1295.
Qi X,Li M W,Xie M,Liu X,Ni M,Shao G,Song C,Yim K Y A,Tao Y,Wong F L,Isobe S,Wong C F,Wong K S,Xu C,Li C,Wang Y,Guan R,Sun F,Fan G,Xiao Z,Zhou F,Phang T H,Liu X,Tong S W,Chan T F,Yiu S M,Tabata S,Wang J,Xu X,Lam H M.2014.Identification of a novel salt tolerance gene in wild soybean by whole-genome sequencing.Nat Commun,5:4340.
Santos C A F,Simon P W.2002.QTL analyses reveal clustered loci for accumulation of major provitamin A carotenes and lycopene in carrot roots.Mol Genet Genomics,268:122-129.
Santos C A F,Simon P W.2006.Heritabilities and minimum gene number estimates of carrot carotenoids.Euphytica,151:79-86.
Simon P W.2000.Domestication,historical development,and modern breeding of carrot.Plant Breed Rev,19:157-190.
Simpson K,Quiroz L F,Rodriguez-Concepción M,Stange C R.2016.Differential contribution of the first two enzymes of the MEP pathway to the supply of metabolic precursors for carotenoid and chlorophyll biosynthesis in carrot(Daucus carota).Front Plant Sci,7:1344.
Umiel N,Gabelman W H.1972.Inheritance of root color and carotenoid synthesis in carrot,Daucus carota L.:orange vs.red.J Am Soc Hort Sci,97:453-460.
Van O J W.2006.JoinM ap 4.0:Software for the calculation of genetic linkage maps in experimental populations.Wageningen,Netherlands:Kyazma B.V.
Vivek B S,Simon P W.1999.Linkage relationships among molecular markers and storage root traits of carro(tDaucus carota L.ssp.sativus).Theor Appl Genet,99:58-64.
Wang H,Ou C G,Zhuang F Y,Ma Z G.2014.The dual role of phytoene synthase genes in carotenogenesis in carrot roots and leaves.Mol Breed,34:2065-2079.
Wang Hui,Ou Cheng-gang,Zhuang Fei-yun,Zhao Zhi-wei,Ma Zhen-guo.2014.Relationship of carotenoid accumulation and transcript of main genes in carotenoid biosynthesis in carrot.Acta Horticulturae Sinica,41(12):2513-2520.(in Chinese)王慧,欧承刚,庄飞云,赵志伟,马振国.2014.胡萝卜中类胡萝卜素积累与主要合成基因转录水平相关性分析.园艺学报,41(12):2513-2520.
Wang L,Wang A,Huang X,Zhao Q,Dong G,Qian Q,Sang T,Han B.2011.Mapping 49 quantitative trait loci at high resolution through sequencing-based genotyping of rice recombinant inbred lines.Theor Appl Genet,122(2):327-340.
Welsch R,Maass D,Voegel T,Dellapenna D,Beyer P.2007.Transcription factor RAP2.2 and its interacting partner SINAT2:stable elements in the carotenogenesis of Arabidopsis leaves.Plant Physiol,145(3):1073-1085.
Xu Z S,Tan H W,Wang F,Hou X L,Xiong A S.2014.Carrot DB:a genomic and transcriptomic database for carrot.Database,doi:10.1093/database/bau096.
Yang J,Hu C C,Ye X Z,Zhu J.2005.QTLNetwork 2.0.Institute of Bioinformatics,Zhejiang University,Hangzhou,China,Available:http://ibi.zju.edu.cn/software/qtlnetwork.
Zhou Z,Zhang C,Zhou Y,Hao Z,Wang Z,Zeng X,Di H,Li M,Zhang D,Yong H,Zhang S,Weng J,Li X.2016.Genetic dissection of maize plant architecture with an ultra-high density bin map based on recombinant inbred lines.BMC Genomics,17:178.