滇龙胆基于叶绿体psbA-trnH、trnL-trnF序列多态性遗传评价
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摘要
目的研究云南各地区包括8个居群116个样品的滇龙胆Gentiana rigescens Franch. ex Hemsl.的遗传多样性,分析滇龙胆遗传分化和遗传结构,评价其遗传变异。方法采用试剂盒按照个体提取滇龙胆的DNA,扩增其叶绿体基因psbA-trnH、trnL-trnF并测序,两个片段拼接后综合分析。结果序列中存在22个变异位点并构成11种单倍体类型,滇龙胆的遗传多样性指数(Hd)为0.737,居群间的遗传分化系数(Gst)为0.37632、基因流(Nm)为0.41,地理区域间Gst为0.19680,Nm为1.02。结论滇龙胆具有较高的遗传多样性,且其遗传变异主要出现在居群间,而2个叶绿体DNA片段在滇龙胆的产地鉴别中意义有限,品质差异的形成可能跟环境因子有较大的关系。
Objective Research on the genetic diversity of Gentiana rigescens Franch. ex Hemsl. including 116 samples from 8 populations in Yunnan, to analyze the genetic differentiation and genetic structure of Gentiana rigescens, and to evaluate its genetic variation. Methods The individual total DNA of G. rigescens was extracted according to DNA extraction kit. The sequence of chloroplast gene psbA-trnH and trnL-trnF was amplified and sequenced, then the two fragments were spliced together for comprehensive analysis. Results There were 22 mutation sites in the sequence and constitute 11 haploid types. The genetic diversity index(Hd) of G. rigescens was 0.737, the genetic differentiation coefficient(Gst) among populations was 0.37632, the gene flow(Nm) was 0.41, and the Gst among geographical regions was 0.19680,Nm was 1.02. Conclusion The G. rigescens had a higher genetic diversity, and its genetic variation mainly occurred among populations. However, two chloroplast DNA fragments were of limited significance in the identification of the origin of G. rigescens, and the formation of quality differences might be mainly related to environmental factors.
Objective Research on the genetic diversity of Gentiana rigescens Franch. ex Hemsl. including 116 samples from 8 populations in Yunnan, to analyze the genetic differentiation and genetic structure of Gentiana rigescens, and to evaluate its genetic variation. Methods The individual total DNA of G. rigescens was extracted according to DNA extraction kit. The sequence of chloroplast gene psbA-trnH and trnL-trnF was amplified and sequenced, then the two fragments were spliced together for comprehensive analysis. Results There were 22 mutation sites in the sequence and constitute 11 haploid types. The genetic diversity index(Hd) of G. rigescens was 0.737, the genetic differentiation coefficient(Gst) among populations was 0.37632, the gene flow(Nm) was 0.41, and the Gst among geographical regions was 0.19680,Nm was 1.02. Conclusion The G. rigescens had a higher genetic diversity, and its genetic variation mainly occurred among populations. However, two chloroplast DNA fragments were of limited significance in the identification of the origin of G. rigescens, and the formation of quality differences might be mainly related to environmental factors.
引文
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[2] 中科院《中国植物志》编委会.中国植物志[M].北京:科学出版社,2013:100.
[3] 国家药典委员会.中国药典(一部)[S].北京:中国医药科技出版社,2010:265.
[4] 褚博文,张霁,李智敏,等.滇龙胆化学成分和药理作用研究进展[J].中国实验方剂学杂志,2016,22(13):213.
[5] 沈磊,谢文菠,王修波,等.滇龙胆不同部位提取物保肝抗炎作用的研究[J].中成药,2017,39(04):701.
[6] 张勇,薛昆鹏,何美,等.固相萃取/超高效液相色谱法测定龙胆泻肝丸中栀子苷、龙胆苦苷与黄芩苷[J].分析测试学报,2013,32(1):122.
[7] 杨雁,邵爱娟,金航,等.云贵高原滇龙胆不同居群形态特征变异研究[J].中草药,2012,43(8):1604.
[8] 孙南君,夏春芳.坚龙胆中化学成分的研究[J].中药通报,1984,9( 1) :33.
[9] 常耀军,曹玉芬,张金梅,等.基于叶绿体DNA分析的辽宁省梨属种质遗传多样性研究[J].园艺学报,2014,41(7):1307.
[10] 杨青松,熊勇,胡琳,等.不同海拔血满草居群cpDNA psbA-trnH序列特征及其遗传结构[J].生物技术,2015,25(04):337.
[11] 赵磊,李智敏,白艳婷,等.滇龙胆地上部分的化学成分研究[J].云南中医学院学报,2009,32(02):27.
[12] 唐荣平,苏汉林,王先宏.滇龙胆草种子萌发及育苗研究[J].种子,2012,31(11):31.
[13] 唐荣平,苏汉林.濒危植物滇龙胆草的生态学、生物学特性研究[J].湖北农业科学,2013,52(14):3364.
[14] 吕伟奇,张霁,左智天,等.滇西北野生滇龙胆根部HPLC指纹图谱研究[J].时珍国医国药,2017,28(01):119.
[15] 周伟,李媛,吴昕怡,等.滇龙胆香叶醇-10-羟化酶基因克隆、生物信息学分析和表达[J].中草药,2017,48(03):546.
[16] 刘小莉,李国栋,王杏婷,等.滇龙胆不同居群rDNA-ITS序列分析[J].时珍国医国药,2016,27(01):104.
[17] Hamilton M B.Four primer pairs for the amplification of chloroplast intergenic regions with intraspecific variation [J].Mol Ecol,1999,8 (3):521
[18] Sang T,Crawford D J,StuessyT F.Chloroplast DNA phylogeny,reticulate evolution,and biogeography of Paeonia (Paeoniaceae) [J] .Am J Bot,1997,84 (8):1120.
[19] Saitou N,Nei M.The neighbor-joining method:A new method for reconstructing phylogenetic trees[J].MoI Biol Evol,1987,4(4):406.
[20] 冯思玲.系统发育树构建方法研究[J].信息技术,2009,(6):38.
[21] Petit RJ,Dtuninil J,Fineschi S,et al.Comparative organization of chloroplast mitochondrial and nuclear diversity in plant populations [J].Mol Ecol,2010,14(3):689.
[22] Soltis D E,Kuzof R K.Discordante between nuclear andchloroplastphylogenies in the Heuchera Group ( Saxifragaceae) [J].Evolution,1995,49:727.
[23] 闫小玲.基于cpDNA单倍型和SSR分析的银杏群体遗传结构和谱系地理学研究[D].浙江大学博士学位论文,2010.
[24] 张得均,高庆波,李福安,等.青海栽培黄管秦艽的叶绿体DNApsbA-trnH核苷酸变异和遗传分析[J].安徽农业科学,2011,39(25):15215.
[25] 张琳,罗智渊,冯丽丽,等.滇龙胆不同居群间品质评价及质量等同性的研究[J].中国药学杂志,2014,49(5):412.
[26] 沈涛,金航,杨涛,等.不同产地野生滇龙胆中主要裂环烯醚萜类成分的含量比较[J].中国实验方剂学杂志,2011,17(13):70.
[27] 李智敏,赵磊,白艳婷,等.不同产地滇龙胆中龙胆苦苷的含量测定[J].云南中医学院学报,2008,31(6):10.