黄芩ITS2条形码数据库构建及其种子的DNA条形码鉴定方法建立

目的:通过构建黄芩核糖体DNA内转录间隔区2(ITS2)条形码数据库,建立黄芩种子DNA条形码鉴定新方法,保障黄芩种子基原准确。方法:收集黄芩对照药材、基原植物、生药材、公共数据库及其常见代用品的ITS2序列,构建黄芩DNA条形码数据库;收集62份市售黄芩种子样品,每份种子获取3~4条ITS2序列,基于BLAST方法、遗传距离法和邻接(NJ)系统发育树法进行物种鉴定。结果:共获得101条黄芩及其代用品的ITS2序列。黄芩的种内序列变异小于种间序列变异,黄芩、甘肃黄芩和粘毛黄芩在NJ系统发育树上分别聚为独立的支,可相互区分。共获得195条黄芩种子的ITS2序列,DNA条形码鉴定结果表明193条为黄芩序列,2条为真菌序列。结论:黄芩ITS2条形码数据库稳定可靠,可满足黄芩种子DNA条形码鉴定的需求。DNA条形码技术可用于黄芩种子的物种鉴定。

Construction of ITS2 Barcode Database of Scutellariae Radix and Establishment of DNA Barcode Identification Method for Its Seeds

Objective: In order to guarantee the seed base of Scutellariae Radix is accurate,an internal transcribed spacer 2 (ITS2) barcode reference library of Scutellariae Radix has been constructed and then a species identification method of Scutellariae Radix seeds based on DNA barcoding technology has been established. Method: ITS2 sequences were obtained from Scutellariae radix reference medicinal materials,original plant,crude medicinal materials,public database,and its substitutes.Totally 62 Scutellariae Radix seed specimens were collected from markets.For each Scutellariae Radix seed specimen,3-4 ITS2 sequences were obtained and then used for species assignment by BLAST method,genetic distance method and neighbor-joining phylogenetic tree method. Result: In total,101 ITS2 sequences of Scutellariae Radix and its substitutes have been obtained.The data analysis result showed that the intraspecific distance of Scutellariae Radix was less than its interspecific distance.The three species of Scutellaria baicalensis,S. rehderiana and S. viscidula were showed separated branch in the neighbor-joining tree.Totally 195 ITS2 sequences have been obtained from Scutellariae Radix seeds,the identification results showed that 193 sequences were Scutellariae Radix and 2 sequences were fungi contaminations. Conclusion: The ITS2 barcode database of Scutellariae Radix is stable and reliable,it can satisfy the demand of DNA barcoding analysis of Scutellariae Radix seeds.