两面针与混伪品及近缘种DNA条形码鉴定研究

Identification of Zanthoxylum nitidum and its adulterants and related species through DNA barcodes

【目的】探讨DNA条形码技术对两面针真伪鉴定的可行性,为两面针种质资源的鉴定提供参考。【方法】对两面针及其常见混伪品和同属近缘种(10种31份样品)进行DNA提取、PCR扩增和双向测序,所得序列经CodonCode Aligner软件处理和人工校对,采用TaxonGap法比较rbcL、trnH-psbA和ITS2等3条序列的物种鉴定力大小;选择适用条形码在不同物种样品范围内进行分析,检查种内种间距离变化情况。【结果】ITS2和trnH-psbA两条序列对10个试验物种的鉴定效率均为100.0%,rbcL在实验物种范围内不存在最小种间遗传距离;ITS2序列在样品鉴定范围由12个物种33个数据扩大为33个物种132个数据时,鉴定成功率从100.0%下降至84.8%,种内变异大于种间变异的比例由71.4%上升至90.1%。【结论】ITS2和trnH-psbA序列可作为两面针真伪鉴定的标准条形码,实际应用中应注意确定物种鉴定样品数及种内充分取样。

[Objective]Identification of Zanthoxylum nitidum and its adulterants and related species through DNA bar- codes was studied to provide references for identifying germplasm resources of Z. nitidum. [ Method ]DNA extracted from 31 samples （10 species） was amplified using PCR and sequenced bidirectionally. The obtained sequences were processed by the software of CodonCode Aligner and checked by proofreaders. Species identification strength of three candidate DNA barcodes of rbcL, trnH-psbA and ITS2 were compared using the tool of TaxonGap. Inter- and within-species variation of the optimal biomarker was examined in two different analytical ranges. [ Result ] Identification efficiency of both ITS2 and trnH-psbA was 100.0%. Regarding rbcL, no minimum interspecifie genetic distance existed within the tested species range. Concerning ITS2, when 33 data of 12 species were expanded to 132 data of 33 species, its identification efficiency was down from 100.0% to 84.8% and the increased rate of intraspecifie variation over interspeeific variation was up from 71.4% to 90.1%. [ Conclusion ]ITS2 and trnH-psbA were recommended as the standard barcodes for identification of Z. nitidum. Identifi- cation sample number and adequate sampling should be taken into account in practical application.