基于DNA条形码的金槐系统发育和变异位点分析  

Phylogenetic and Mutation Points Analysis of Sophora japonica cv.jinhuai Based on DNA Barcoding Sequences

作　　者：何爱芳 傅鹏[1] 陈恒宇 蒋柳晶 冯晓晴 HE Ai-fang;FU Peng;CHEN Heng-yu;JIANG Liu-jing;FENG Xiao-qing(Guangxi University of Chinese Medicine,Nanning 530200,China)

机构地区：[1]广西中医药大学,南宁530200

出　　处：《中国实验方剂学杂志》2022年第12期183-191,共9页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：广西中医药大学2020年硕士研究生科研创新项目(YCSY2020097);广西壮瑶药重点实验室开放课题项目(GXZYZZ2020A-12)。

摘　　要：目的:对来自不同居群的金槐资源的核糖体内转录间隔区2(ITS2)和叶绿体基因片段psbA-trnH、rbcL、matK进行系统发育分析,探索不同地理来源的金槐资源的遗传多样性。方法:用聚合酶链式反应(PCR)扩增获得金槐ITS2、psbA-trnH、rbcL和matK的核酸序列,用邻接法(NJ)构建系统发育树,用Kimura 2-Parameter(K2P)模型分析遗传距离,用MEGA和BIOEDIT软件进行多重比对分析。结果:ITS2序列为278~279 bp;psbA-trnH为289 bp;rbcL为673 bp;matK为786~792 bp。ITS2和psbA-trnH都存在3个变异位点,rbcL没有变异位点,matK有13个变异位点。采用ITS2序列构建的系统发育树将金槐样品资源分为2个大类群,百宝实生树聚为一类,其余25份资源聚为一类。对于psbA-trnH序列,28份金槐资源的PCR扩增成功率为100%。psbA-trnH序列聚类结果显示,28份金槐资源可分为3个大的类群。庙头嫁接/实生树、枧塘嫁接树、文桥嫁接树、咸水实生树和大圩嫁接树组成一个类群;绍水实生树单独组成一个类群;其余21份资源组成一个类群。基于matK序列建立的发育树可将26份金槐资源分为2个类群,其中咸水实生树单独为一个分支,其余25份金槐资源聚为一支。对金槐的rbcL序列的聚类结果无法将28份资源区分开。基于ITS2+psbA-trnH+rbcL+matK组合序列构建的系统发育树可将金槐资源分为4个大类群,祁阳实生/嫁接树、道县实生树,庙头嫁接/实生树、永岁嫁接树、绍水实生树、石塘实生树、咸水实生树、枧塘实生树、文桥嫁接树、秧塘嫁接树、湘漓实生树聚为一支;文桥实生树聚为一支;大圩实生树聚为一支;其他金槐资源聚为一支。结论:系统发育和变异位点分析为金槐资源的进化研究、道地性评价奠定了理论基础,变异位点的分析可用于相关资源的鉴定。结果也表明将不同基因片段组合对植物进行鉴定,效果更佳。Objective:To conduct phylogenetic analysis of internal transcribed spacer 2(ITS2)and chloroplast gene segments including psbA-trnH,rbcL,and matK of Sophora japonica cv.jinhuai resource samples from different geographical sources,and to explore the genetic diversity of S.japonica cv.jinhuai.Method:Polymerase chain reaction(PCR)method was used to amplify the nucleic acid sequences of ITS2,psbA-trnH,rbcL,and matK of S.japonica cv.jinhuai.Neighbor joining(NJ)method was used to construct phylogenetic trees,and Kimura 2-Parameter(K2P)model was used to calculate the genetic distance of different samples.MEGA and BIOEDIT softwares were applied for mutiple alignment and analysis of ITS2,psbA-trnH,rbcL,and matK sequences of S.japonica cv.jinhuai.Result:The lengths of ITS2 sequence were 278-279 bp.The lengths of psbA-trnH were 289 bp.The lengths of rbcL sequence were 673 bp.The lengths of matK sequences were 786-792 bp.There were 3 mutation points in ITS2 and psbA-trnH,no mutation point in rbcL,and 13 mutation points in matK.The samples of S.japonica cv.jinhuai were clustered into two groups based on the phylogenetic tree constructed by ITS2 sequences.The sample of seedling tree in Baibao was clustered into one group,while the other 25 samples were clustered into another group.For the psbA-trnH sequence,the success rate of PCR amplification of 28 samples of S.japonica cv.jinhuai was 100%.The 28 samples of S.japonica cv.jinhuai were clustered into three groups based on the clustering results of psbA-trnH sequence.The sample of seedling tree in Shaoshui was clustered into one group.The five samples of grafting tree and seedling tree in Miaotou,grafting trees in Jiantang,Wenqiao,and Daxu,and seeding tree in Xianshui were clustered into one group.The other 21 samples were clustered into another group.The 26 samples of S.japonica cv.jinhuai were clustered into two groups based on the phylogenetic tree constructed by matK sequences.The sample of seedling tree in Xianshui was clustered into one group,while the other 25 samples wer

关 键 词：金槐 内转录间隔区(2ITS2) PSBA-TRNH RBCL MATK 系统发育 聚合酶链式反应(PCR) 

分 类 号：R284.2[医药卫生—中药学] R289[医药卫生—中医学]