基于SSR分子标记的野生山胡椒(Lindera glauca)核心种质的构建
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摘要
为深入发掘和筛选中国野生山胡椒种质资源,本研究利用18对SSR引物(5叶绿体SSR和13核SSR),以基本涵盖中国大陆自然分布区的22个自然群体共303份野生山胡椒种质为材料,进行SSR基因分型分析,并绘制出原始种质的亲缘关系图。结果表明,18对引物共检测到Na=88,Ne为1.18～3.76,He为0.14～0.74,Ho为0～0.58,I为0.30～1.52,以及PIC为0.13～0.70。STRUCTURE分析和UPGMA聚类的结果都显示出22个自然群体适合被划分为三个亚类群。利用等位基因最大化(M策略)和遗传距离最大化(SAGD)法分别至少要在25%(71份种质)、15%取样比例下(45份种质)构建的核心种质才能够在所有遗传多样性指标上代表原始种质的遗传多样性(t检验不显著),并符合核心种质构建的标准。结合核心种质构建工作的同类研究,确定SAGD法(15%取样比例)是较适宜于构建山胡椒核心种质的方法。基于nSSR标记,SAGD法15%比例下构建的核心种质在Na、Ne和I指标参数上的保留率分别达到了原始种质的93.24%、104.67%和100.85%;基于cpSSR标记,与之对应的保留率分别为85.14%、107.45%和108.82%。通过此法筛选出的45份核心种质材料能够一定程度地代表中国整个野生山胡椒种质资源的遗传多样性。研究结果可为其他对非作物型木本植物的核心种质构建,尤其是兼有无性与有性繁殖的物种提供参考。
In order to further explore and screen the germplasm resources of wild Lindera glauca in China, 18 pairs of SSR primers(5 chloroplast SSR and 13 nuclear SSR) were used in this study to analyze the SSR genotyping of 303 wild Lindera glauca germplasms from 22 natural populations that basically covered the natural distribution areas in mainland China and the genetic relationship diagram of original germplasm was drawn. The results showed that 18 pairs of primers detected Na=88. Ne was 1.18～3.76 and He was 0.14～0.74. Ho was 0～0.58 and I was 0.30～1.52, as well as PIC was 0.13～0.70. The results of STRUCTURE analysis and UPGMA clustering showed that 22 natural populations were suitable for being divided into three subgroups. The core germplasm constructed by using allele maximization(M strategy) and genetic distance maximization(SAGD) methodsat least25%(71 germplasm) and 15%(45 germplasm) sample rate, respectively could represent genetic diversity of the original germplasmon all genetic diversity indicators(t test was not significant), and conformed to the standard ofcore germplasm construction. Compared with other similar research of core germplasm, the SAGD(15% sampling ratio) was a suitable method for constructing L. glauca core germplasm. Based on n SSR markers, the retention rate of Na, Ne, and I index parameters of core germplasm constructed by 15% proportion of SAGD reached 93.24%,104.67% and 100.85% of the original germplasm, respectively. Based on cpSSR markers, the corresponding retention rate were 85.14%, 107.45% and 108.82%, respectively. And the 45 core germplasm materials screened by this method could represent the genetic diversity of the whole wild L. glauca germplasm resources in China to a certain extent. The research results herein would provide references for other core germplasm construction of non-crop woody plants, especially for species that have both asexual and sexual reproduction.
In order to further explore and screen the germplasm resources of wild Lindera glauca in China, 18 pairs of SSR primers(5 chloroplast SSR and 13 nuclear SSR) were used in this study to analyze the SSR genotyping of 303 wild Lindera glauca germplasms from 22 natural populations that basically covered the natural distribution areas in mainland China and the genetic relationship diagram of original germplasm was drawn. The results showed that 18 pairs of primers detected Na=88. Ne was 1.18～3.76 and He was 0.14～0.74. Ho was 0～0.58 and I was 0.30～1.52, as well as PIC was 0.13～0.70. The results of STRUCTURE analysis and UPGMA clustering showed that 22 natural populations were suitable for being divided into three subgroups. The core germplasm constructed by using allele maximization(M strategy) and genetic distance maximization(SAGD) methodsat least25%(71 germplasm) and 15%(45 germplasm) sample rate, respectively could represent genetic diversity of the original germplasmon all genetic diversity indicators(t test was not significant), and conformed to the standard ofcore germplasm construction. Compared with other similar research of core germplasm, the SAGD(15% sampling ratio) was a suitable method for constructing L. glauca core germplasm. Based on n SSR markers, the retention rate of Na, Ne, and I index parameters of core germplasm constructed by 15% proportion of SAGD reached 93.24%,104.67% and 100.85% of the original germplasm, respectively. Based on cpSSR markers, the corresponding retention rate were 85.14%, 107.45% and 108.82%, respectively. And the 45 core germplasm materials screened by this method could represent the genetic diversity of the whole wild L. glauca germplasm resources in China to a certain extent. The research results herein would provide references for other core germplasm construction of non-crop woody plants, especially for species that have both asexual and sexual reproduction.
引文
Belaj A.,Dominguez-García M.C.,Atienza S.G.,Urdíroz N.M.,Raúl D.R.,Satovic Z.,Martín A.,Kilian A.,Trujillo I.,Valpuesta V.,and Carmen D.R.,2012,Developing a core collection of olive(Olea europaea L.)based on molecular markers(DArTs,SSRs,SNPs)and agronomic traits,Tree Genet.Genomes,8(2):365-378
Ben-Ayed R.,Sans-Grout C.,Moreau F.,Grati-Kamoun N.,and Rebai A.,2014,Genetic similarity among Tunisian olive cultivars and two unknown feral olive trees estimated through SSR markers,Biochem.Genet.,52(5-6):258-268
Brown A.H.D.,1989,Core collection:a practical approach to genetic resources management,Genome,31(2):818-824
Bu H.D.,and Zhang B.B.,2010,Advances in research on core collection of fruit tree germplasm resources,Beifang Yuanyi(Northern Horticulture),(4):211-213(卜海东,张冰冰,2010,果树核心种质研究进展,北方园艺,(4):211-213)
Cipriani G.,Spadotto A.,Jurman I.,Gaspero G.D.,Crespan M.,Meneghetti S.,Frare E.,Vignani R.,Cresti M.,Morgante M.,Pezzotti M.,Pe E.,Policriti A.,and Testolin R.,2010,The SSR-based molecular profile of 1005 grapevine(Vitis vinifera L.)accessions uncovers new synonymy and parentages,and reveals a large admixture amongst varieties of different geographic origin,Theor.Appl.Genet.,121(8):1569-1585
Daniel G.,Olivier L.,Enrico P.,and Grossniklaus U.,2001,Developmental genetics of gametophytic apomixes,Trends Genet.,17(10):597-604
Doyle J.J.,and Doyle J.L.,1987,A rapid DNA isolation procedure for small quantities of fresh leaf tissue,Phytochem.Bull.,19(1):11-15
Dupont Y.L.,2002,Evolution of apomixis as a strategy of colonization in the dioecious species Lindera glauca(Lauraceae),Population Ecology,44(3):293-297
Earl D.A.,and vonHoldt B.M.,2012,STRUCTURE HAR-VESTER:a website and program for visualizing structure output and implementing the evanno method,Conserv.Genet.Resour.,4(2):359-361
Ergül A.,Türkoglu M.,and Soylemezoglu G.,2004,Genetic identification of Amasya(Vitis vinifera L.Cvs.)genotypes based on AFLP markers,Biotechnol.,Biotec.,Eq.,18(3):39-43
Frankel O.H.,and Brown A.H.D.,1984,Plant genetic resources today:a critical appraisal,In:Holden J.H.W.,and Williams J.T.(eds.),Crop genetic resources:conservation and evaluation,George Allan and Unwin,London,UK,pp.249-257
Guo D.L.,Liu C.H.,Zhang J.Y.,and Zhang G.H.,2012,Construction of grape core collections,Zhongguo Nongye Kexue(Scientia Agricultura Sinica),45(6):1135-1143(郭大龙,刘崇怀,张君玉,张国海,2012,葡萄核心种质的构建,中国农业科学,45(6):1135-1143)
Hu J.,Zhu J.,and Xu H.M.,2000,Methods of constructing core collections by stepwise clustering with three sampling strategies based on the genotypic values of crops,Theor.Appl.Genet.101(1):264-268
Kim K.H.,Moon E.,Ha K.,Ha S.K.,Sun W.S.,Kim H.K.,Kim S.Y.,Choi S.U.,and Lee K.R.,2014,Bioactive lignan constituents from the twigs of Lindera glauca,Chem.Pharm.Bull.,62(11):1136-1140
Kim K.W.,Chung H.K.,Cho G.T.,Ma K.H.,Chandrabalan D.,Gwag J.G.,Kim T.S.,Cho E.G.,and Park Y.J.,2007,PowerCore:a program applying the advanced M strategy with a heuristic search for establishing core sets,Bioinformatics,23(16):2155-2162
Liu J.,Liao K.,Mansur N.,Cao Q.,Jiang Z.B.,and Jia Y.,2015,Core-germplasm construction of apricot collections in south of Xinjiang by ISSR molecular markers,Guoshu Xuebao(Journal of Fruit Science),32(3):374-384(刘娟,廖康,曼苏尔·那斯尔,曹倩,江振斌,贾杨,2015,利用ISSR分子标记构建南疆杏种质资源核心种质,果树学报,32(3):374-384)
Liu K.,and Muse S.V.,2005,PowerMarker:an integrated analysis environment for genetic marker analysis,Bioinformatics,21(9):2128-2129
Liu L.D.,Chen J.D.,and Lan S.G.,1982,Chemical composition test and research applications of Lindera galuca fruits,Zhiwu Xuebao(Journal of Integrative Plant Biology),24(3):252-258(刘立鼎,陈京达,兰胜桂,1982,山胡椒果实化学成分研究应用试验,植物学报,24(3):252-258)
Liu Y.Y.,Mei H.X.,Du Z.W.,Wu K.,Zhen Y.Z.,Cui X.H.,and Zhen L.,2017,Construction of core collection of sesame based on phenotype and molecular markers,Zhongguo Nongye Kexue(Scientia Agricultura Sinica),50(13):2433-2441(刘艳阳,梅鸿献,杜振伟,武轲,郑永战,崔向华,郑磊,2017,基于表型和SSR分子标记构建芝麻核心种质,中国农业科学,50(13):2433-2441)
Mahar K.S.,Rana T.S.,and Ranade S.A.,2011,Molecular analyses of genetic variability in soap nut(Sapindusmukorossi Gaertn.),Ind.Crops Products,34(1):1111-1118
Mao J.,Liu X.L.,Su H.S.,Lu X.,Lin X.Q.,Cai Q.,and Fan Y.H.,2016,Constructing core collection of Saccharum arundinaceum L.based on phenotype and molecular markers,Zhiwu Yichuan Ziyuan Xuebao(Journal of Plant Genetic Resources),17(4):607-615(毛钧,刘新龙,苏火生,陆鑫,林秀琴,蔡青,范源洪,2016,基于表型与分子数据的斑茅核心种质构建,植物遗传资源学报,17(4):607-615)
Marzouki H.,Nasri N.,Jouaud B.,Bonnet C.,Khaldi A.,Bouzid S.,and Fady B.,2009,Population genetic structure of Laurus nobilis L.inferred from transferred nuclear microsatellites,Silvae Genetica,58(5):270-276
Niu J.,Hou X.Y.,Fang C.L.,An J.Y.,Ha D.L.,Qiu L.,Ju Y.X.,Zhao H.Y.,Du W.Z.,Qi J.,Zhang Z.X.,Liu G.N.,and Lin S.Z.,2015,Transcriptome analysis of distinct Lindera glauca tissues revealed the differences in the Uni Genes related to terpenoid biosynthesis,Gene,559(1):22-30
Nybom H.,2004,Comparison of different nuclear DNA markers for estimating intraspecific genetic diversity in plants,Molecular Ecology,13:1143-1155
Odong T.,Heerwaarden J.,Jansen J.,Hintumc T.J.L.,and Eeuwijka F.A.,2011,Statistical techniques for defining reference sets of accessions and microsatellite markers,Crop Sci.,51(6):2401-2411
Pelsy F.,Hocquigny S.,Moncada X.,Barbeau G.,Forget D.,Hinrichsen P.,and Merdinoglu D.,2010,An extensive study of the genetic diversity within seven French wine grape variety collections,Theor.Appl.Genet.120(6):1219-1231
Pritchard J.K.,Stephens M.,and Donnelly P.,2000,Inference of population structure using multilocus genotype data:dominant markers and null alleles,Genetics,7(4):574-578
Qi J.,Xiong B.,Zhang Z.X.,and Zhen A.G.,and Pu F.G.,2016,Diversity analysis on fatty acid composition of Lindera glauca fruit from different provenances,Zhongguo Liangyou Xuebao(Journal of the Chinese Cereals and Oils Association),31(12):72-78(齐季,熊彪,张志翔,甄爱国,蒲发光,2016,不同种源山胡椒果实脂肪酸组分多样性分析,中国粮油学报,31(12):72-78)
Smouse P.E.,Whitehead M.R.,and Peakall R.,2015,An informational diversity framework,illustrated with sexually deceptive orchids in early stages of speciation,Molecular E-cology Resources,15(6):1375-1384
Sreekumar V.B.,and Renuka C.,2006,Assessment of genetic diversity in Calamus thwaitesii BECC(Arecaceae)using RAPDmarkers,Biochemi.Syst.Ecol.,34(5):397-405
Sudheer P.D.V.,Singh S.,Mastan S.G.,Patel J.,and Reddy M.P.,2009,Molecular characterization and identification of markers for toxic and non-toxic varieties of Jatropha curcas L.using RAPD,AFLP and SSR markers,Molecular Biology Reports,36(6):1357-1364
Suh W.S.,Kim K.H.,Kim H.K.,Choi S.U.,and Lee K.R.,2015,Three new lignan derivatives from Lindera glauca(Siebold et Zucc)Blume,Helvetica Chimica Acta,98(8):1087-1094
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Xiong B.,Dong S.B.,Qi J.,Zhang L.M.,Ha D.L.,Ju Y.X.,and Zhang Z.X.,2016,Development and characterization of microsatellite loci for Lindera glauca(Lauraceae),Appli.Plant Sci.,4(11):1600088
Xiong B.,Qi J.,Dong S.B.,Zhang L.M.,Zhang Z.X.,Ha D.L.,and Ju Y.X.,2016,Superior individual selection of linderaglauca based on fruit and seed traits,Dongbei Linye Daxue Xuebao(Journal of Northeast Forestry University),44(5):10-14(熊彪,齐季,董树斌,张莉梅,张志翔,哈登龙,琚煜熙,2016,基于种实性状的山胡椒优良单株选择,东北林业大学学报,44(5):10-14)
Yang H.B.,Zhang R.,Wang B.S.,Xu Z.Y.,and Zhou Z.C.,2017,Construction of core collection of Schimasuperba based on SSR molecular markers,Linye Kexue(Scientia Silvae Sinicae),53(6):37-46(杨汉波,张蕊,王帮顺,徐肇友,周志春,2017,基于SSR标记的木荷核心种质构建,林业科学,53(6):37-46)
Zhang C.Y.,Chen X.S.,Zhang Y.M.,Yuan Z.H.,Liu Z.C.,Wang Y.L.,and Lin Q.,2009,A method for constructing core collection of Malus sieversii using molecular markers,Zhongguo Nongye Kexue(Scientia Agricultura Sinica),42(2):597-604(张春雨,陈学森,张艳敏,苑兆和,刘遵春,王延龄,林群,2009,采用分子标记构建新疆野苹果核心种质的方法,中国农业科学,42(2):597-604)
Ben-Ayed R.,Sans-Grout C.,Moreau F.,Grati-Kamoun N.,and Rebai A.,2014,Genetic similarity among Tunisian olive cultivars and two unknown feral olive trees estimated through SSR markers,Biochem.Genet.,52(5-6):258-268
Brown A.H.D.,1989,Core collection:a practical approach to genetic resources management,Genome,31(2):818-824
Bu H.D.,and Zhang B.B.,2010,Advances in research on core collection of fruit tree germplasm resources,Beifang Yuanyi(Northern Horticulture),(4):211-213(卜海东,张冰冰,2010,果树核心种质研究进展,北方园艺,(4):211-213)
Cipriani G.,Spadotto A.,Jurman I.,Gaspero G.D.,Crespan M.,Meneghetti S.,Frare E.,Vignani R.,Cresti M.,Morgante M.,Pezzotti M.,Pe E.,Policriti A.,and Testolin R.,2010,The SSR-based molecular profile of 1005 grapevine(Vitis vinifera L.)accessions uncovers new synonymy and parentages,and reveals a large admixture amongst varieties of different geographic origin,Theor.Appl.Genet.,121(8):1569-1585
Daniel G.,Olivier L.,Enrico P.,and Grossniklaus U.,2001,Developmental genetics of gametophytic apomixes,Trends Genet.,17(10):597-604
Doyle J.J.,and Doyle J.L.,1987,A rapid DNA isolation procedure for small quantities of fresh leaf tissue,Phytochem.Bull.,19(1):11-15
Dupont Y.L.,2002,Evolution of apomixis as a strategy of colonization in the dioecious species Lindera glauca(Lauraceae),Population Ecology,44(3):293-297
Earl D.A.,and vonHoldt B.M.,2012,STRUCTURE HAR-VESTER:a website and program for visualizing structure output and implementing the evanno method,Conserv.Genet.Resour.,4(2):359-361
Ergül A.,Türkoglu M.,and Soylemezoglu G.,2004,Genetic identification of Amasya(Vitis vinifera L.Cvs.)genotypes based on AFLP markers,Biotechnol.,Biotec.,Eq.,18(3):39-43
Frankel O.H.,and Brown A.H.D.,1984,Plant genetic resources today:a critical appraisal,In:Holden J.H.W.,and Williams J.T.(eds.),Crop genetic resources:conservation and evaluation,George Allan and Unwin,London,UK,pp.249-257
Guo D.L.,Liu C.H.,Zhang J.Y.,and Zhang G.H.,2012,Construction of grape core collections,Zhongguo Nongye Kexue(Scientia Agricultura Sinica),45(6):1135-1143(郭大龙,刘崇怀,张君玉,张国海,2012,葡萄核心种质的构建,中国农业科学,45(6):1135-1143)
Hu J.,Zhu J.,and Xu H.M.,2000,Methods of constructing core collections by stepwise clustering with three sampling strategies based on the genotypic values of crops,Theor.Appl.Genet.101(1):264-268
Kim K.H.,Moon E.,Ha K.,Ha S.K.,Sun W.S.,Kim H.K.,Kim S.Y.,Choi S.U.,and Lee K.R.,2014,Bioactive lignan constituents from the twigs of Lindera glauca,Chem.Pharm.Bull.,62(11):1136-1140
Kim K.W.,Chung H.K.,Cho G.T.,Ma K.H.,Chandrabalan D.,Gwag J.G.,Kim T.S.,Cho E.G.,and Park Y.J.,2007,PowerCore:a program applying the advanced M strategy with a heuristic search for establishing core sets,Bioinformatics,23(16):2155-2162
Liu J.,Liao K.,Mansur N.,Cao Q.,Jiang Z.B.,and Jia Y.,2015,Core-germplasm construction of apricot collections in south of Xinjiang by ISSR molecular markers,Guoshu Xuebao(Journal of Fruit Science),32(3):374-384(刘娟,廖康,曼苏尔·那斯尔,曹倩,江振斌,贾杨,2015,利用ISSR分子标记构建南疆杏种质资源核心种质,果树学报,32(3):374-384)
Liu K.,and Muse S.V.,2005,PowerMarker:an integrated analysis environment for genetic marker analysis,Bioinformatics,21(9):2128-2129
Liu L.D.,Chen J.D.,and Lan S.G.,1982,Chemical composition test and research applications of Lindera galuca fruits,Zhiwu Xuebao(Journal of Integrative Plant Biology),24(3):252-258(刘立鼎,陈京达,兰胜桂,1982,山胡椒果实化学成分研究应用试验,植物学报,24(3):252-258)
Liu Y.Y.,Mei H.X.,Du Z.W.,Wu K.,Zhen Y.Z.,Cui X.H.,and Zhen L.,2017,Construction of core collection of sesame based on phenotype and molecular markers,Zhongguo Nongye Kexue(Scientia Agricultura Sinica),50(13):2433-2441(刘艳阳,梅鸿献,杜振伟,武轲,郑永战,崔向华,郑磊,2017,基于表型和SSR分子标记构建芝麻核心种质,中国农业科学,50(13):2433-2441)
Mahar K.S.,Rana T.S.,and Ranade S.A.,2011,Molecular analyses of genetic variability in soap nut(Sapindusmukorossi Gaertn.),Ind.Crops Products,34(1):1111-1118
Mao J.,Liu X.L.,Su H.S.,Lu X.,Lin X.Q.,Cai Q.,and Fan Y.H.,2016,Constructing core collection of Saccharum arundinaceum L.based on phenotype and molecular markers,Zhiwu Yichuan Ziyuan Xuebao(Journal of Plant Genetic Resources),17(4):607-615(毛钧,刘新龙,苏火生,陆鑫,林秀琴,蔡青,范源洪,2016,基于表型与分子数据的斑茅核心种质构建,植物遗传资源学报,17(4):607-615)
Marzouki H.,Nasri N.,Jouaud B.,Bonnet C.,Khaldi A.,Bouzid S.,and Fady B.,2009,Population genetic structure of Laurus nobilis L.inferred from transferred nuclear microsatellites,Silvae Genetica,58(5):270-276
Niu J.,Hou X.Y.,Fang C.L.,An J.Y.,Ha D.L.,Qiu L.,Ju Y.X.,Zhao H.Y.,Du W.Z.,Qi J.,Zhang Z.X.,Liu G.N.,and Lin S.Z.,2015,Transcriptome analysis of distinct Lindera glauca tissues revealed the differences in the Uni Genes related to terpenoid biosynthesis,Gene,559(1):22-30
Nybom H.,2004,Comparison of different nuclear DNA markers for estimating intraspecific genetic diversity in plants,Molecular Ecology,13:1143-1155
Odong T.,Heerwaarden J.,Jansen J.,Hintumc T.J.L.,and Eeuwijka F.A.,2011,Statistical techniques for defining reference sets of accessions and microsatellite markers,Crop Sci.,51(6):2401-2411
Pelsy F.,Hocquigny S.,Moncada X.,Barbeau G.,Forget D.,Hinrichsen P.,and Merdinoglu D.,2010,An extensive study of the genetic diversity within seven French wine grape variety collections,Theor.Appl.Genet.120(6):1219-1231
Pritchard J.K.,Stephens M.,and Donnelly P.,2000,Inference of population structure using multilocus genotype data:dominant markers and null alleles,Genetics,7(4):574-578
Qi J.,Xiong B.,Zhang Z.X.,and Zhen A.G.,and Pu F.G.,2016,Diversity analysis on fatty acid composition of Lindera glauca fruit from different provenances,Zhongguo Liangyou Xuebao(Journal of the Chinese Cereals and Oils Association),31(12):72-78(齐季,熊彪,张志翔,甄爱国,蒲发光,2016,不同种源山胡椒果实脂肪酸组分多样性分析,中国粮油学报,31(12):72-78)
Smouse P.E.,Whitehead M.R.,and Peakall R.,2015,An informational diversity framework,illustrated with sexually deceptive orchids in early stages of speciation,Molecular E-cology Resources,15(6):1375-1384
Sreekumar V.B.,and Renuka C.,2006,Assessment of genetic diversity in Calamus thwaitesii BECC(Arecaceae)using RAPDmarkers,Biochemi.Syst.Ecol.,34(5):397-405
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