基于DNA条形码的龙葵系统发育及变异位点分析

目的对来自主要龙葵产区龙葵种质资源的转录间隔区(internal transcribed spacer,ITS)和trn H-psb A基因间隔区序列进行系统发育学分析,为龙葵资源的合理利用和道地性评价提供理论基础。方法用聚合酶链式反应(polymerase chain reaction,PCR)的方法扩增龙葵ITS区和trn H-psb A的DNA序列,并与美国国家生物技术信息中心(NCBI)数据库进行比对获取ITS1+ITS2和trn H-psb A的DNA序列。用邻接法(Neighbor joining,NJ)和最大简约法(maximum parsimony,MP)构建系统发育树,用Kimura two-parameter(K2-P)模型分析遗传距离,用Clustal X和DNAman软件进行多重比对。结果 17个龙葵样品的ITS1序列长度均为230 bp,ITS2序列长度均为206 bp,trn H-psb A序列长度为446 bp或447 bp,3个序列分别存在7个、2个和3个变异位点。系统发育的方法将中国龙葵种质资源聚为3个类群,这些类群与其所处的纬度存在一定的相关性。遗传距离的分析结果显示来自广东梅州的样品与来自北京和河北等地的样品存在最大的遗传距离。结论系统发育和变异位点的分析为中国龙葵资源的利用和进化研究,以及龙葵资源的道地性评价提供了理论基础,变异位点的分析还可应用于相关种质资源的鉴定。

Phylogenetic and mutation point analysis of DNA barcoding sequences in Solanum nigrum

Objective In this study, phylogenetic analysis was used to compare the ITSs and trnH-psbA sequences of 17Solanum nigrum samples, providing the theoretic foundation to utilize their resources and evaluate their genuineness. Methods PCR method was used to amplify the region of ITS and trnH-psbA, and the seqeucens of ITS1+ITS2 and trnH-psbA were obtained after the amplified fragment sequences were blasted in NCBI database. The Neighbor joining(NJ) and maximum parsimony(MP) method were used to construct phylogenetic trees and Kimura two-parameter(K2-P) model was used to calculate the genetic distance of different samples. Clustal X and DNAman softwares were applied for multi-alignment of ITS1, ITS2, and trnH-psbA sequences from different samples. Results The lengths of ITS1 and ITS2 sequences from 17 samples were 230 and 206 bp, respectively, and trnH-psbA sequences were 446 or 447 bp. ITS1, ITS2, and trnH-psbA had seven, two, and three mutation points, respectively. These 17 samples were clustered to three latitude-dependent groups based on both ITS1+ITS2 and trnH-psbA sequences. Conclusion Phylogenetic and mutation point analysis will provide the theoretic foundation to utilize the resources of Chinese S. nigrum, investigate their evolution, and evaluate their genuineness. The results of mutation point will also be used in the identification of related S. nigrum resources.