摘要目的 评价3种不同的分子标记技术对植物属级的鉴定能力以选出最适合在中毒现场快速鉴定有毒植物的分子技术. 方法 选取毛茛科、大戟科共18种19份有毒植物叶片样品,用改良十六烷基三甲基溴化铵法提取基因组DNA,分别使用随机扩增多态性DNA标记(RAPD)、简单重复序列间标记(ISSR)和DNA条形码技术进行物种鉴定(属级),使用NYSTS、SPSS、PAUP、MEGA软件进行聚类分析,比较3种DNA分子标记技术的准确性、可靠性、时效性和可操作性. 结果 准确性:RAPD技术未能鉴定样品的属级,ISSR技术和DNA条形码技术的鉴定准确率分别为68％和100％.可靠性:RAPD技术与ISSR技术主观影响率分别为44％、26％,条带重复率分别为47％、45％；DNA条形码技术trnH-psbA片段扩增成功率与测序成功率均为100％.时效性:从总DNA提取开始至得到最终鉴定结果,RAPD、ISSR、DNA条形码技术耗时分别为8.5、9.0、42.2 h.可操作性:RAPD与ISSR技术的鉴定工作可在普通实验室完成,DNA条形码技术的鉴定工作则需要特殊的测序设备. 结论 初步认为DNA条形码技术是比较适用于突发有毒植物中毒事件中快速病因判定的分子鉴定技术.

abstractsObjective To evaluate the efficiency of the three molecular marker techniques for identification of plant genus in order to select the most suitable molecular technique to rapidly identify a poisonous plant at the scene of the poisoning accident.Methods Nineteen samples of leaves in 18 species of common poisonous plants of Ranunculaceae and Euphorbiaceaec were selected.Genomic DNA of these samples were extracted using improved hexadecyltrimethylammonium bromide (CTAB) method,species identification (genus level) were performed using random amplified polymorphic DNA (RAPD),intersimple sequences repeats (ISSR) and DNA barcoding,cluster analysis were performed using NYSTS,SPSS,PAUP,and MEGA software.Accuracy,reliability,timeliness,and operability were compared between the 3 molecular marker techniques.Results Accuracy:genus level of samples failed to be identified using RAPD ; and the accuracy rates in genus level of identification using ISSR and DNA barcoding were 68％ and 100％,respectively.Reliability:the subjective influence rates and the repetitive rates in RAPD and ISSR were 47％ and 26％ as well as 47％ and 45％,respectively; the success rates of amplification and sequencing of trnH-psbA fragment using DNA barcoding both were 100％.Timeliness:the time from extraction of DNA to accomplishment of identification in the 3 molecular marker techniques were 8.5,9.0,and 42.2 h,respectively.Operability:species identification using RAPD and ISSR could be done in a common laboratory; special sequencing equipment was needed when using DNA barcoding technique in the identification of poisonous plants.Conclusion The preliminary study results show that DNA barcoding is a molecular identification technique more suitable for rapid determination of the poisonous plant at the scene of the sudden poisoning accident.