多基原民族药岩陀DNA条形码鉴定

Identification of Multi-source Ethnodrug Rodgersiae Rhizoma by DNA Barcoding Technology

目的:采用ITS,ITS2,rbcL,matK和psbA-trnH这5种热门DNA条形码序列对多基原民族药岩陀的鉴别能力进行评价,筛选出适合该物种鉴定的DNA条形码。方法:通过比较各序列聚合酶链式反应(PCR)扩增成功率及测序成功率,计算种内种间遗传距离,运用Wilcoxon非参数秩和检验进行遗传差异分析,邻接(NJ)法构建系统发育树以及采用Blast 1和NJ建树方法评价不同序列对物种的鉴定成功率等策略的实施,综合评价上述5种DNA候选条形码在多基原岩陀植物中的鉴别效果。结果:ITS序列扩增成功率及测序成功率分别为100%和96.61%;ITS2序列扩增成功率及测序成功率分别为100%和98.31%;psbA-trnH序列扩增成功率及测序成功率均为100%;rbcL序列扩增成功率及测序成功率均为100%,matK扩增成功率及测序成功率均为98.31%。psbA-trnH序列拥有最高的种内及种间遗传变异,且种内最大变异的平均值小于种间最小变异的平均值。系统发育树构建结果显示,psbA-trnH序列能区分3种不同来源的岩陀药用植物,而ITS,ITS2,rbcL和matK序列聚类效果较差。psbA-trnH序列鉴定成功率均为100%,而ITS,ITS2,matK和rbcL序列对物种鉴定成功率均低于50%。结论:psbA-trnH较其他序列有明显优势,因此推荐psbA-trnH序列作为理想的DNA条形码,用于多基原民族药岩陀的鉴定研究。

Objective:Five popular DNA barcoding sequences,namely ITS,ITS2,rbcL,matK and psbA-trnH,were employed to evaluate the identification efficiency of multi-source ethnodrug Rodgersiae Rhizoma,and the most suitable sequence was then screened out.Method:Efficiency of polymerase chain reaction(PCR)amplification,success rate of sequencing,intra-and inter-specific distances calculated by rank sum test,phylogenetic tree constructed with neighbor-joining(NJ)method and identification efficiency assessed by Blast 1 and NJ method were adopted in this study.Result:Efficiency of PCR amplification and success rate of sequencing for ITS,ITS2,psbA-trnH,rbcL and matK were 100%,96.61%,100%,98.31%,100%,100%,100%,100%,98.31%and 98.31%,respectively.Intra-and inter-specific genetic distances and identification achievement rate for psbA-trnH were the highest among the five candidate sequences.Besides,the average coalescent depth was less than the smallest interspecific distance for psbA-trnH.Phylogenetic tree also illustrated that Rodgersiae Rhizoma could be distinguished based on psbA-trnH.Conclusion:According to the findings,psbA-trnH was superior to other DNA barcodes.Therefore,psbA-trnH was recommended as the ideal DNA barcode for the identification of multi-source ethnodrug Rodgersiae Rhizoma.