利用重测序–BSA分析鉴定金柑油胞发育相关基因
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摘要
为了鉴定与金柑油胞发育相关的基因,用多油胞的罗纹金柑[Fortunella japonica(Thunb.)Swingle,母本]和少油胞的滑皮金柑(Fortunella crassifolia Swingle ‘Huapi’,父本)进行杂交构建了F_1代分离群体,从159个F1代单株中挑选出高油胞密度和低油胞密度的单株各30株,分别构建两个极端性状的DNA混池用于基因组重测序。利用重测序数据结合混合分组分析法(Bulked Segregant Analysis,BSA)进行遗传关联性分析,将控制油胞数量性状的位点定位在第9条染色体的25 980 001～29 160 001 bp的区域内。该区域包含511个有效的SNP,分布在301个基因位点上。同源性功能分析后获得11个与细胞程序性死亡、细胞壁功能和结构及细胞伸长和膨大等生理过程相关的候选基因(Ciclev10005243m.g、Ciclev10005288m.g、Ciclev10005338m.g、Ciclev10005441m.g、Ciclev10006448m.g、Ciclev10005804m.g、Ciclev10004719m.g、Ciclev10005888m.g、Ciclev10006502m.g、Ciclev10005197m.g和Ciclev10004431m.g),这些基因可能在柑橘油胞的发育过程中起重要作用。
The density of oil gland is an important factor to determine the content of essential oil in citrus. To understand the genetic control of oil gland development,a segregation population of 159 F_1 individuals was constructed by crossing Luowen kumquat[Fortunella japonica(Thunb.)Swingle]with Huapi kumquat(F. crassifolia Swingle‘Huapi'),a low oil gland density mutation variety. Thirty individuals of each extreme phenotype group with either high-density or low-density oil glands were chosen and their DNA were pooled to construct two DNA pools for genomic re-sequencing to identify candidate genes associated with oil gland development. The results of Bulked Segregant Analysis(BSA)indicated that the 25 980 001–29 160 001 bp region of chromosome 9 is highly correlated to the density of oil glands. Through analysis of recombination frequency of SNP,511 SNP loci distributed in 301 genes were found to be possibly associated with the density of oil glands. The homology function analysis suggested that 11 of these genes(Ciclev10005243 m.g,Ciclev10005288 m.g,Ciclev10005338 m.g,Ciclev10005441 m.g,Ciclev10006448 m.g,Ciclev10005804 m.g,Ciclev10004719 m.g,Ciclev10005888 m.g,Ciclev10006502 m.g,Ciclev10005197 m.g and Ciclev10004431 m.g)involving in cell programmed death,the function and construction of cell wall,and cell elongation and expansion may be the important genetic factors controlling the development of oil gland.
The density of oil gland is an important factor to determine the content of essential oil in citrus. To understand the genetic control of oil gland development,a segregation population of 159 F_1 individuals was constructed by crossing Luowen kumquat[Fortunella japonica(Thunb.)Swingle]with Huapi kumquat(F. crassifolia Swingle‘Huapi'),a low oil gland density mutation variety. Thirty individuals of each extreme phenotype group with either high-density or low-density oil glands were chosen and their DNA were pooled to construct two DNA pools for genomic re-sequencing to identify candidate genes associated with oil gland development. The results of Bulked Segregant Analysis(BSA)indicated that the 25 980 001–29 160 001 bp region of chromosome 9 is highly correlated to the density of oil glands. Through analysis of recombination frequency of SNP,511 SNP loci distributed in 301 genes were found to be possibly associated with the density of oil glands. The homology function analysis suggested that 11 of these genes(Ciclev10005243 m.g,Ciclev10005288 m.g,Ciclev10005338 m.g,Ciclev10005441 m.g,Ciclev10006448 m.g,Ciclev10005804 m.g,Ciclev10004719 m.g,Ciclev10005888 m.g,Ciclev10006502 m.g,Ciclev10005197 m.g and Ciclev10004431 m.g)involving in cell programmed death,the function and construction of cell wall,and cell elongation and expansion may be the important genetic factors controlling the development of oil gland.
引文
Chavez D J,Chaparro J X.2011.Identification of markers linked to seedlessness in Citrus kinokuni hort.ex Tanaka and its progeny using bulked segregant analysis.HortScience,46(5):693-697.
Chen Y,Wu H.2010.Programmed cell death involved in the schizolysigenous formation of the secretory cavity in Citrus sinensis L.(Osbeck).Chinese Science Bulletin,55(20):2160-2168.
Clement Y,Sarah G,Holtz Y,Homa F,Pointet S,Contreras S,Nabholz B,Sabot F,Saune L,Ardisson M,Bacilieri R,Besnard G,Berger A,Cardi C,De Bellis F,Fouet O,Jourda C,Khadari B,Lanaud C,Leroy T,Pot D,Sauvage C,Scarcelli N,Tregear J,Vigouroux Y,Yahiaoui N,Ruiz M,Santoni S,Labouisse J P,Pham J L,David J,Glemin S.2017.Evolutionary forces affecting synonymous variations in plant genomes.PLoS Genetics,13(5):e1006799.
Cummins I,Landrum M,Steel P G,Edwards R.2007.Structure activity studies with xenobiotic substrates using carboxylesterases isolated from Arabidopsis thaliana.Phytochemistry,68(6):811-818.
Dong W,Wu D,Li G,Wu D,Wang Z.2018.Next-generation sequencing from bulked segregant analysis identifies a dwarfism gene in watermelon.Scientic Report,8(1):2908.
Gao Heng-jin.2016.Study on genetic diversity of citrus germplasm based on the molecular markers[M.D.Dissertation].Chongqing:Southwest University.(in Chinese)高恒锦.2016.基于分子标记柑橘遗传多样性研究[硕士论文].重庆:西南大学.
Kayam G,Brand Y,Faigenboim-Doron A,Patil A,Hedvat I,Hovav R.2017.Fine-mapping the branching habit trait in cultivated peanut by combining bulked segregant analysis and high-throughput sequencing.Frontiers in Plant Science,doi:10.3389/fpls.2017.00467.
Knight T G,Klieber A,Sedgley M.2001.The relationship between oil gland and fruit development in Washington Navel orange(Citrus sinensis L.Osbeck).Annual Botany London,88(6):1039-1047.
Lei Tian-gang,Zou Xiu-ping,He Yong-rui,Yao Li-xiao,Peng Ai-hong,Li Qiang,Xu Lan-zhen,Chen Shan-chun.2015.Genetic background analysis of several guizhou local citrus varieties based on SSR markers.Journal of Southwest University(Natural Science Edition),37(11):30-35.(in Chinese)雷天刚,邹修平,何永睿,姚利晓,彭爱红,李强,许兰珍,陈善春.2015.贵州部分柑橘地方品种遗传背景的SSR分析.西南大学学报(自然科学版),37(11):30-35.
Liang S J,Wang H Y,Yang M,Wu H.2009.Sequential actions of pectinases and cellulases during secretory cavity formation in Citrus fruits.Trees,23(1):19-27.
Lionetti V,Fabri E,Caroli M D,Hansen A R,Willats W G T,Piro G,Bellincampi D.2017.Three pectin methyl esterase inhibitors protect cell wall integrity for immunity to Botrytis.Plant Physiology,173(3):1844-1863.
Liu J L,Park C H,He F,Nagano M,Wang M,Bellizzi M,Zhang K,Zeng X S,Liu W D,Ning Y S,Kawano Y,Wang G L.2015.The RhoGAPSPIN6 associates with SPL11 and OsRac1 and negatively regulates programmed cell death and innate immunity in rice.PLoS Pathogens,11(2):e1004629.
Liu P,Liang S,Yao N,Wu H.2012.Programmed cell death of secretory cavity cells in fruits of Citrus grandis cv.Tomentosa is associated with activation of caspase 3-like protease.Trees,26(6):1821-1835.
Luro F,Venturini N,Costantino G,Paolini J,Ollitrault P,Costa J.2012.Genetic and chemical diversity of citron(Citrus medica L.)based on nuclear and cytoplasmic markers and leaf essential oil composition.Phytochemistry,77:186-196.
Marshall S D,Putterill J J,Plummer K M,Newcomb R D.2003.The carboxylesterase gene family from Arabidopsis thaliana.Journal of Molecular Evolution,57(5):487-500.
Michelmore R W,Paran I,Kesseli R V.1991.Identification of markers linked to disease-resistance genes by bulked segregant analysis:a rapid method to detect markers in specific genomic regions by using segregating populations.Proceedings of the National Academy of Sciences of the United States of America,88(21):9828-9832.
Mindrebo J T,Nartey C M,Seto Y,Burkart M D,Noel J P.2016.Unveiling the functional diversity of the alpha/beta hydrolase superfamily in the plant kingdom.Current Opinion in Structural Biology,41:233-246.
Morlais I,Severson D W.2001.Identification of a polymorphic mucin-like gene expressed in the midgut of the mosquito,Aedes aegypti,using an integrated bulked segregant and differential display analysis.Genetics,158(3):1125-1136.
Orth T,Reumann S,Zhang X,Fan J,Wenzel D,Quan S,Hu J.2007.The PEROXIN11 protein family controls peroxisome proliferation in Arabidopsis.Plant Cell,19(1):333-350.
Pan Y J,Liu L,Lin Y C,Zu Y G,Li L P,Tang Z H.2016.Ethylene antagonizes salt-induced growth retardation and cell death process via transcriptional controlling of ethylene-,BAG-and senescence-associated genes in Arabidopsis.Frontiers in Plant Science,7:696.
Rafiei M.,Rajaei H.2007.Structural and developmental studies on oil producing reproductive organs in lime(Citrus aurantifolia Swingle).Journal of Biological Sciences,7(6):848-853.
Shahidi F,Zhong Y.2005.Citrus oils and essences//Shadihi F.Bailey’s industrial oil and fat products.USA:Wiley-Interscience.
Thomson W W,Plattaloia K A,Endress A G.1976.Ultrastructure of oil gland development in leaf of Citrus sinensis L.Botanical Gazette,137(4):330-340.
Tranchida P Q,Bonaccorsi I,Dugo P,Mondello L,Dugo G.2012.Analysis of Citrus essential oils:state of the art and future perspectives.A review.Flavour and Fragrance Journal,27(2):98-123.
van der Lee T,Robold A,Testa A,van’t Klooster J W,Govers F.2001.Mapping of avirulence genes in Phytophthora infestans with amplified fragment length polymorphism markers selected by bulked segregant analysis.Genetics,157(3):949-956.
Verica J A,He Z H.2002.The cell wall-associated kinase(WAK)and WAK-like kinase gene family.Plant Physiology,129(2):455-459.
Voo S S,Grimes H D,Lange B M.2012.Assessing the biosynthetic capabilities of secretory glands in Citrus peel.Plant Physiology,159(1):81-94.
Wang Jiong.2017.Citrus linkage map construction based on COS marker and photosynthetic characteristics research of the mapping population[M.D.Dissertation].Chongqing:Southwest University.(in Chinese)王炯.2017.基于COS Marker构建柑橘连锁图谱及作图群体的光合特性研究[硕士论文].重庆:西南大学.
Wang Jiong,Gong Gui-zhi,Peng Zhu-chun,Li Yi-bing,Wang Yan-jie,Hong Qi-bin.2017.Genetic and phylogenetic relationships among Citrus and its close and distant relatives based on COS Marker.Scientia Agricultura Sinica,50(2):320-331.(in Chinese)王炯,龚桂芝,彭祝春,李一兵,王艳杰,洪棋斌.2017.基于COS Marker分析柑橘属及其近缘、远缘属植物的遗传与进化.中国农业科学,50(2):320-331.
Wang X,Xu Y,Zhang S,Cao L,Huang Y,Cheng J,Wu G,Tian S,Chen C,Liu Y.2017.Genomic analyses of primitive,wild and cultivated citrus provide insights into asexual reproduction.Nature Genetics,49(5):765-772.
Xia Y,Yin S J,Zhang K L,Shi X T,Lian C L,Zhang H S,Hu Z B,Shen Z G.2018.OsWAK11,a rice wall-associated kinase,regulates Cu detoxification by alteration the immobilization of Cu in cell walls.Environmental and Experimental Botany,150:99-105.
Xue H B,Shi T,Wang F F,Zhou H K,Yang J,Wang L,Wang S,Su Y L,Zhang Z,Qiao Y S,Li X G.2017.Interval mapping for red/green skin color in Asian pears using a modified QTL-seq method.Horticulture Research,doi:10.1038/hortres.2017.53.
Yang Ke.1985.Chang’an kumquat new strain-Huapi kumquat.Zhejiang Citrus,(2):38-39.(in Chinese)杨科.1985.长安金柑新品系--滑皮金柑.浙江柑橘,(2):38-39.
Ye Yin-min.1983.Germplasm resources of Chinese kumquat.Crop Germplasm,(4):2-5.(in Chinese)叶荫民.1983.中国金柑种质资源.作物品种资源,(4):2-5.
Yu Hui-wen.2018.Investigation on the genetic diversity and leaf slow-greening mutation of pummelo[Ph.D.Dissertation].Wuhan:Huazhong Agricultral University.(in Chinese)余惠文.2018.柚遗传多样性及叶片渐绿性状突变研究[博士论文].武汉:华中农业大学.
Zhang Lian-feng,He Jian,Feng Yan,Liu Li,Guo Qi-gao,Liang Guo-lu.2006.Phylogenetic relationships among Fortunella and its relatives as revealed by SSR markers.Journal of Fruit Science,23(3):335-338.(in Chinese)张连峰,何建,冯焱,刘利,郭启高,梁国鲁.2006.金柑属及其近缘属植物亲缘关系的SSR分析.果树学报,23(3):335-338.
Zheng P,Bai M,Chen Y,Liu P W,Gao L,Liang S J,Wu H.2014.Programmed cell death of secretory cavity cells of citrus fruits is associated with Ca2+accumulation in the nucleus.Trees,28(4):1137-1144.
Chen Y,Wu H.2010.Programmed cell death involved in the schizolysigenous formation of the secretory cavity in Citrus sinensis L.(Osbeck).Chinese Science Bulletin,55(20):2160-2168.
Clement Y,Sarah G,Holtz Y,Homa F,Pointet S,Contreras S,Nabholz B,Sabot F,Saune L,Ardisson M,Bacilieri R,Besnard G,Berger A,Cardi C,De Bellis F,Fouet O,Jourda C,Khadari B,Lanaud C,Leroy T,Pot D,Sauvage C,Scarcelli N,Tregear J,Vigouroux Y,Yahiaoui N,Ruiz M,Santoni S,Labouisse J P,Pham J L,David J,Glemin S.2017.Evolutionary forces affecting synonymous variations in plant genomes.PLoS Genetics,13(5):e1006799.
Cummins I,Landrum M,Steel P G,Edwards R.2007.Structure activity studies with xenobiotic substrates using carboxylesterases isolated from Arabidopsis thaliana.Phytochemistry,68(6):811-818.
Dong W,Wu D,Li G,Wu D,Wang Z.2018.Next-generation sequencing from bulked segregant analysis identifies a dwarfism gene in watermelon.Scientic Report,8(1):2908.
Gao Heng-jin.2016.Study on genetic diversity of citrus germplasm based on the molecular markers[M.D.Dissertation].Chongqing:Southwest University.(in Chinese)高恒锦.2016.基于分子标记柑橘遗传多样性研究[硕士论文].重庆:西南大学.
Kayam G,Brand Y,Faigenboim-Doron A,Patil A,Hedvat I,Hovav R.2017.Fine-mapping the branching habit trait in cultivated peanut by combining bulked segregant analysis and high-throughput sequencing.Frontiers in Plant Science,doi:10.3389/fpls.2017.00467.
Knight T G,Klieber A,Sedgley M.2001.The relationship between oil gland and fruit development in Washington Navel orange(Citrus sinensis L.Osbeck).Annual Botany London,88(6):1039-1047.
Lei Tian-gang,Zou Xiu-ping,He Yong-rui,Yao Li-xiao,Peng Ai-hong,Li Qiang,Xu Lan-zhen,Chen Shan-chun.2015.Genetic background analysis of several guizhou local citrus varieties based on SSR markers.Journal of Southwest University(Natural Science Edition),37(11):30-35.(in Chinese)雷天刚,邹修平,何永睿,姚利晓,彭爱红,李强,许兰珍,陈善春.2015.贵州部分柑橘地方品种遗传背景的SSR分析.西南大学学报(自然科学版),37(11):30-35.
Liang S J,Wang H Y,Yang M,Wu H.2009.Sequential actions of pectinases and cellulases during secretory cavity formation in Citrus fruits.Trees,23(1):19-27.
Lionetti V,Fabri E,Caroli M D,Hansen A R,Willats W G T,Piro G,Bellincampi D.2017.Three pectin methyl esterase inhibitors protect cell wall integrity for immunity to Botrytis.Plant Physiology,173(3):1844-1863.
Liu J L,Park C H,He F,Nagano M,Wang M,Bellizzi M,Zhang K,Zeng X S,Liu W D,Ning Y S,Kawano Y,Wang G L.2015.The RhoGAPSPIN6 associates with SPL11 and OsRac1 and negatively regulates programmed cell death and innate immunity in rice.PLoS Pathogens,11(2):e1004629.
Liu P,Liang S,Yao N,Wu H.2012.Programmed cell death of secretory cavity cells in fruits of Citrus grandis cv.Tomentosa is associated with activation of caspase 3-like protease.Trees,26(6):1821-1835.
Luro F,Venturini N,Costantino G,Paolini J,Ollitrault P,Costa J.2012.Genetic and chemical diversity of citron(Citrus medica L.)based on nuclear and cytoplasmic markers and leaf essential oil composition.Phytochemistry,77:186-196.
Marshall S D,Putterill J J,Plummer K M,Newcomb R D.2003.The carboxylesterase gene family from Arabidopsis thaliana.Journal of Molecular Evolution,57(5):487-500.
Michelmore R W,Paran I,Kesseli R V.1991.Identification of markers linked to disease-resistance genes by bulked segregant analysis:a rapid method to detect markers in specific genomic regions by using segregating populations.Proceedings of the National Academy of Sciences of the United States of America,88(21):9828-9832.
Mindrebo J T,Nartey C M,Seto Y,Burkart M D,Noel J P.2016.Unveiling the functional diversity of the alpha/beta hydrolase superfamily in the plant kingdom.Current Opinion in Structural Biology,41:233-246.
Morlais I,Severson D W.2001.Identification of a polymorphic mucin-like gene expressed in the midgut of the mosquito,Aedes aegypti,using an integrated bulked segregant and differential display analysis.Genetics,158(3):1125-1136.
Orth T,Reumann S,Zhang X,Fan J,Wenzel D,Quan S,Hu J.2007.The PEROXIN11 protein family controls peroxisome proliferation in Arabidopsis.Plant Cell,19(1):333-350.
Pan Y J,Liu L,Lin Y C,Zu Y G,Li L P,Tang Z H.2016.Ethylene antagonizes salt-induced growth retardation and cell death process via transcriptional controlling of ethylene-,BAG-and senescence-associated genes in Arabidopsis.Frontiers in Plant Science,7:696.
Rafiei M.,Rajaei H.2007.Structural and developmental studies on oil producing reproductive organs in lime(Citrus aurantifolia Swingle).Journal of Biological Sciences,7(6):848-853.
Shahidi F,Zhong Y.2005.Citrus oils and essences//Shadihi F.Bailey’s industrial oil and fat products.USA:Wiley-Interscience.
Thomson W W,Plattaloia K A,Endress A G.1976.Ultrastructure of oil gland development in leaf of Citrus sinensis L.Botanical Gazette,137(4):330-340.
Tranchida P Q,Bonaccorsi I,Dugo P,Mondello L,Dugo G.2012.Analysis of Citrus essential oils:state of the art and future perspectives.A review.Flavour and Fragrance Journal,27(2):98-123.
van der Lee T,Robold A,Testa A,van’t Klooster J W,Govers F.2001.Mapping of avirulence genes in Phytophthora infestans with amplified fragment length polymorphism markers selected by bulked segregant analysis.Genetics,157(3):949-956.
Verica J A,He Z H.2002.The cell wall-associated kinase(WAK)and WAK-like kinase gene family.Plant Physiology,129(2):455-459.
Voo S S,Grimes H D,Lange B M.2012.Assessing the biosynthetic capabilities of secretory glands in Citrus peel.Plant Physiology,159(1):81-94.
Wang Jiong.2017.Citrus linkage map construction based on COS marker and photosynthetic characteristics research of the mapping population[M.D.Dissertation].Chongqing:Southwest University.(in Chinese)王炯.2017.基于COS Marker构建柑橘连锁图谱及作图群体的光合特性研究[硕士论文].重庆:西南大学.
Wang Jiong,Gong Gui-zhi,Peng Zhu-chun,Li Yi-bing,Wang Yan-jie,Hong Qi-bin.2017.Genetic and phylogenetic relationships among Citrus and its close and distant relatives based on COS Marker.Scientia Agricultura Sinica,50(2):320-331.(in Chinese)王炯,龚桂芝,彭祝春,李一兵,王艳杰,洪棋斌.2017.基于COS Marker分析柑橘属及其近缘、远缘属植物的遗传与进化.中国农业科学,50(2):320-331.
Wang X,Xu Y,Zhang S,Cao L,Huang Y,Cheng J,Wu G,Tian S,Chen C,Liu Y.2017.Genomic analyses of primitive,wild and cultivated citrus provide insights into asexual reproduction.Nature Genetics,49(5):765-772.
Xia Y,Yin S J,Zhang K L,Shi X T,Lian C L,Zhang H S,Hu Z B,Shen Z G.2018.OsWAK11,a rice wall-associated kinase,regulates Cu detoxification by alteration the immobilization of Cu in cell walls.Environmental and Experimental Botany,150:99-105.
Xue H B,Shi T,Wang F F,Zhou H K,Yang J,Wang L,Wang S,Su Y L,Zhang Z,Qiao Y S,Li X G.2017.Interval mapping for red/green skin color in Asian pears using a modified QTL-seq method.Horticulture Research,doi:10.1038/hortres.2017.53.
Yang Ke.1985.Chang’an kumquat new strain-Huapi kumquat.Zhejiang Citrus,(2):38-39.(in Chinese)杨科.1985.长安金柑新品系--滑皮金柑.浙江柑橘,(2):38-39.
Ye Yin-min.1983.Germplasm resources of Chinese kumquat.Crop Germplasm,(4):2-5.(in Chinese)叶荫民.1983.中国金柑种质资源.作物品种资源,(4):2-5.
Yu Hui-wen.2018.Investigation on the genetic diversity and leaf slow-greening mutation of pummelo[Ph.D.Dissertation].Wuhan:Huazhong Agricultral University.(in Chinese)余惠文.2018.柚遗传多样性及叶片渐绿性状突变研究[博士论文].武汉:华中农业大学.
Zhang Lian-feng,He Jian,Feng Yan,Liu Li,Guo Qi-gao,Liang Guo-lu.2006.Phylogenetic relationships among Fortunella and its relatives as revealed by SSR markers.Journal of Fruit Science,23(3):335-338.(in Chinese)张连峰,何建,冯焱,刘利,郭启高,梁国鲁.2006.金柑属及其近缘属植物亲缘关系的SSR分析.果树学报,23(3):335-338.
Zheng P,Bai M,Chen Y,Liu P W,Gao L,Liang S J,Wu H.2014.Programmed cell death of secretory cavity cells of citrus fruits is associated with Ca2+accumulation in the nucleus.Trees,28(4):1137-1144.