基于核基因ITS及叶绿体psbA-trnH和trnS-trnG基因怀地黄栽培起源探讨
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摘要
目的从野生型驯化为栽培品种角度分析地黄Rehmannia glutinosa的遗传多样性,探讨河南焦作地区怀地黄的栽培起源。方法对地黄野生自然居群,栽培品种及其近缘物种ITS、psb A-trn H和trn S-trn G序列进行扩增和测序,分析比较中药材地黄野生自然居群,栽培品种间各序列的单倍型类型和核苷酸多态性,构建Neighbor-Joining(NJ)分子系统发育树。结果单倍型分析结果显示,地黄野生居群单倍型类型数量(ITS 6个,psb A-trn H 8个,trn S-trn G 9个)明显高于怀地黄栽培品种单倍型类型数量(ITS 3个,psb A-trn H 2个,trn S-trn G 3个)。地黄野生居群的单倍型多样性和核苷酸多样性均远高于怀地黄栽培品种间,在3个基因联合系统树上,地黄野生居群和栽培品种所有样品与茄叶地黄聚在一支,支持率为90%;23个栽培品种(29个样本)与来自温县的10号地黄野生居群聚为一支,支持率为78%。结论地黄野生型驯化为栽培品种过程发生了明显的遗传瓶颈,导致现存怀地黄栽培品种遗传基础狭窄,遗传多样性降低。怀地黄栽培品种可能起源于河南温县区域的地黄野生群体。
Objective In order to understand the genetic diversity of cultivated and wild Rehmannia glutinosa, and to unravel the origin of cultivated R. glutinosa. Methods The sequences of nuclear gene ITS and chloroplast gene psb A-trnH, trnS-trnG in cultivar and wild population of R. glutinosa were amplified and sequenced. Haplotype(gene) diversity and nucleotide polymorphism of three genes from wild and cultivars of R. glutinosa were analyzed and compared in this study. Phylogenetic tree was constructed based on combined three genes using Bayesian inference(BI) methods. Results Analysis of sequences indicated that the number of haplotype(ITS, 6; psb A-trnH, 8; trnS-trnG, 9) in wild population of R. glutinosa was obviously higher than the number of Haplotype(ITS, 3; psb A-trnH, 3; trnS-trnG, 3) in cultivars of R. glutinosa. Haplotype diversity and nucleotide polymorphism of wild population of R. glutinosa were far higher than that of cultivars of R. glutinosa. The NJ tree(combined three genes data) indicated that all cultivated and wild population of R. glutinosa, and R. solanifolia form a monophyletic clade [Posterior probability(PP) = 90%]. Twenty-three cultivars of R.glutinosa(including 29 samples) were clustered with Wenxian wild populations(PP = 78%). Conclusion The results implied very low genetic diversity existed in cultivars of R. glutinosa induced by the severe genetic bottleneck during the process of domestication of wild R. glutinosa, which resulted in the narrow genetic basis of the existing cultivars and decreased genetic diversity. Furthermore, it appeared that wild populations in Wenxian-Henan area were involved in the origin of cultivars of R. glutinosa.
Objective In order to understand the genetic diversity of cultivated and wild Rehmannia glutinosa, and to unravel the origin of cultivated R. glutinosa. Methods The sequences of nuclear gene ITS and chloroplast gene psb A-trnH, trnS-trnG in cultivar and wild population of R. glutinosa were amplified and sequenced. Haplotype(gene) diversity and nucleotide polymorphism of three genes from wild and cultivars of R. glutinosa were analyzed and compared in this study. Phylogenetic tree was constructed based on combined three genes using Bayesian inference(BI) methods. Results Analysis of sequences indicated that the number of haplotype(ITS, 6; psb A-trnH, 8; trnS-trnG, 9) in wild population of R. glutinosa was obviously higher than the number of Haplotype(ITS, 3; psb A-trnH, 3; trnS-trnG, 3) in cultivars of R. glutinosa. Haplotype diversity and nucleotide polymorphism of wild population of R. glutinosa were far higher than that of cultivars of R. glutinosa. The NJ tree(combined three genes data) indicated that all cultivated and wild population of R. glutinosa, and R. solanifolia form a monophyletic clade [Posterior probability(PP) = 90%]. Twenty-three cultivars of R.glutinosa(including 29 samples) were clustered with Wenxian wild populations(PP = 78%). Conclusion The results implied very low genetic diversity existed in cultivars of R. glutinosa induced by the severe genetic bottleneck during the process of domestication of wild R. glutinosa, which resulted in the narrow genetic basis of the existing cultivars and decreased genetic diversity. Furthermore, it appeared that wild populations in Wenxian-Henan area were involved in the origin of cultivars of R. glutinosa.
引文
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[2]王太霞,李景原,胡正海.地黄的形态结构与化学成分研究进展[J].中草药,2004,35(5):585-587.
[3]丁自勉.地黄[M].北京:中国中医药出版社,2001.
[4]中国药典[S].一部.2015.
[5]赵燏黄.国产地黄类生药学的研究[J].药学学报,1953,1(1):41.
[6]温学森,杨世林,魏建和,等.地黄栽培历史及其品种考证[J].中草药,2002,33(10):946-949.
[7]温学森,李先恩,赵华英,等.地黄常见种质的染色体观察[J].中草药,2005,36(1):124-125.
[8]李宏庆.地黄属分类学与系统学研究[D].上海:华东师范大学,2005.
[9]夏至.地黄属和崖白菜属的分子系统学研究—兼论地黄属和崖白菜属的科级系统位置[D].北京:中国科学院植物研究所,2009.
[10]夏至,李家美.地黄属及其近缘属的药用植物资源调查研究[J].商丘师范学院学报,2009,25(12):96-98.
[11]赵楠.地黄的遗传多样性研究[D].上海:华东师范大学,2007.
[12]赵楠,李宏庆.地黄居群遗传多样性的ISSR分析[J].河南科学,2009,27(11):1386-1391.
[13]王云生,黄宏文,王瑛.作物驯化的遗传学研究及其在大豆育种中的应用[J].植物学通报,2008,25(2):221-229.
[14]刘彦飞,梁东,罗桓,等.地黄的化学成分研究[J].中草药,2014,45(1):16-22.
[15]邢洁,徐为人,刘鹏,等.栀子和地黄环烯醚萜类成分抗炎作用的虚拟评价[J].中草药,2009,40(6):930-935.
[16]陈京荔,黄璐琦,邵爱娟,等.地黄不同品种的RAPD分析[J].中国中药杂志,2002,27(7):505-508.
[17]Qi JJ,Li X E,Song J Y,et al.Genetic Relationships among Rehmannia glutinosa cultivars and varieties[J].Planta Med,2008,74(15):1846-1852.
[18]周延清,景建洲,李振勇,等.利用RAPD和ISSR分子标记分析地黄种质遗传多样性[J].遗传,2004,26(6):922-928.
[19]刘春艳.地黄属植物遗传多样性和遗传结构的研究[D].西安:西北大学.2015.
[20]张重义,李改玲,牛苗苗,等.连作地黄的生理生态响应与品质评价[J].中国中药杂志,2011,36(9):1133-1136.
[21]夏至,王璐静,黄勇,等.地黄属植物DNA条形码鉴定及地黄栽培起源研究[J].中草药,2016,47(4):648-654.
[22]Wendel J F,Schnabel A S,Seelanan T.Bidirectional inter locus concerted evolution following allopolyploid speciation in cotton(Gossypium)[J].Proc Natl Acad SciUSA,1995,92:280-284.
[23]Shaw J,Lickey E B,Beck J T,et al.The tortoise and the hare II:Relative utility of 21 noncoding chloroplast DNA sequences for phylogenetic analysis[J].Am J Bot,2005,92(1):142-166.
[24]Xia Z,Wang Y Z,Smith J F.Familial placement and relationships of Rehmannia and Triaenophora(Scrophulariaceae sensu lato)inferred from five different gene regions[J].Am J Bot,2009,96(2):519-530.
[25]Thompson J D,Gibson T J,Plewniak F,et al.The Clustal X windows interface:Flexible strategies for multiple sequence alignment aided by quality analysis tools[J].Nucl Acids Res,1997,25(25):4876-4882.
[26]Hall T A.BIOEDIT A user-friendly biological sequence alignment editor and analysis program for Windows95/98/NT[J].Nucl Acids Symp Ser,1999,41(41):95-98.
[27]Xia X,Xie Z.DAMBE:Software package for data analysis in molecular biology and evolution[J].J Hered,2001,30(7):371-373.
[28]Librado P,Rozas J.Dna SP v5:A software for comprehensive analysis of DNA polymorphism data[J].Bioinformatics,2009,25(11):1451-1452.
[29]Ronquist F,Huelsenbeck J P.Mr Bayes 3:Bayesian phyloge-netic inference under mixed models[J].Bioinformatics 2003,19:1572-1574.
[30]Posada D,Crandall K A.Modeltest:Testing the model of DNA substitution[J].Bioinformatics,1998,14:817-818.
[31]Tanksley S D,Mccouch S R.Seed banks and molecular maps:Unlocking genetic potential from the wild[J].Science,1997,277:1063-1066.
[32]曹元宇.本草经[M].上海:上海科学技术出版社,1987.
[33]刘文泰(明).本草品精要[M].北京:人民卫生出版社,1982.
[34]李时珍(明).本草纲目(上册)[M].北京:人民卫生出版社,1982.
[35]陈川.药用植物玄参的栽培起源、亲缘地理及东亚玄参系统发育研究[D].杭州:浙江大学,2011.