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【目的】DNA条形码技术已成为生物分类鉴定的有力工具。DNA条形码技术的相关问题,如物种种内和种间的遗传距离出现重叠区域,将直接影响到物种鉴定的准确性。我们应用DNA条形码试剂盒检测技术来快速、准确地鉴定口岸截获的检疫性大小蠹属种类。【方法】针对大小蠹昆虫设计引物以提高PCR扩增效率。运用自主研发的基因条码分析软件找出基因片段上区分每个物种的多态位点规律,作为该物种的鉴定特征并建立数据库,应用于物种鉴定。【结果】使用针对大小蠹属昆虫设计的引物成功扩增出325 bp的COI基因片段。将大小蠹属12种昆虫的COI基因片段上的核苷酸诊断位点的组合作为物种的鉴定特征,可以准确地区分近似种。通过比对植物检疫鉴定系统数据库里的鉴定特征,将6个大小蠹属的未知样品成功鉴定到种(核苷酸序列一致性为100%),与形态鉴定结果一致。【结论】结果表明DNA条形码试剂盒检测技术可以准确鉴定大小蠹属的种类。该检测技术可以应用于其他经济重要性有害生物的检测鉴定。
【Objective】 DNA barcoding technology has become a powerful tool for taxonomic identification. The problems associated with DNA barcoding, such as overlapping genetic distances within and between species, will have a direct impact on the accuracy of species identification. We applied the DNA barcode kit detection technique to quickly and accurately identify the quarantine species Dendroctonus species quarantined at the port. 【Method】 Primers were designed to enhance the PCR amplification efficiency. Using self-developed gene barcode analysis software to identify the genetic segments of each species polymorphism site rules, as the identification characteristics of the species and the establishment of a database used in species identification. 【Result】 A 325 bp COI gene fragment was successfully amplified using primers designed for the genus Dendrolimus. The combination of the nucleotide diagnostic sites on the COI gene fragments of 12 species of the genus Dendrantheus can be used as identification features of the species to accurately distinguish the approximate species. By comparing the identification features in the database of phytosanitary systems, 6 unknown samples of Dendrolimus were successfully identified (the nucleotide sequence identity was 100%), which was consistent with the results of morphological identification. 【Conclusion】 The results show that the DNA barcode kit detection technology can accurately identify the species of Dendroctonus. The detection technology can be applied to other economically important pests detection and identification.