应用DNA条形码技术鉴定口岸截获的国内未见分布蚊种

DNA Barcoding Identification of China Non-Recorded Mosquito Species Intercepted at the Port

[目的]利用DNA条形码技术对口岸截获的形态特征破坏严重的未知蚊类进行种类鉴定。[方法]提取从荷兰进口的废纸中截获的一头形态严重破坏的蚊类的基因组DNA。采用无脊椎动物线粒体细胞色素氧化酶I(COI)的通用引物LCO1490和HCO2198扩增目的片段。然后纯化、克隆测序、序列分析、与BOLD中的序列进行比对。[结果]该未知蚊类的DNA条形码序列与BOLD中环带林蚊(Sylvicola cinctus)的序列相似性超过99.8%;并与原产地专家交流确认截获的蚊种为环带林蚊。经科技查新,确认为首次截获的国内未见分布蚊种。[结论]DNA条形码能克服传统物种鉴定依靠完整的外部形态的局限,有效提高外来物种的种类鉴别能力,大大缩短检测周期,降低外来物种对我国生态和公共卫生安全的危害。

[Objective] DNA barcoding techniques were utilized to identify the unknown mosquito species intercepted from Zhongshan port, whose morphology was damaged too badly to be identified. [Method] Genomic DNA was extracted from the mosquito which was found in the waste paper imported from Netherlands and the morphological characteristics were seriously damaged. Target fragments were amplifi ed with the universal primers LCO1490 and HCO2198 of the invertebrate mitochondrial cytochrome oxidase I （COI）, PCR products were purified, cloned and sequenced. The sequences were blasted in the BOLD. [Results] The sequence of the DNA barcode fragment from the unknown mosquito was more than 99.8% identical to the published sequence ofSylvicola cinctus in BOLD. The results were confi rmed by the expert from Europe where S.cinctus is a common species. S.cinctus is never recorded in China and it is thefi rst time to intercepte this species at the port according to the result of ‘science and technology novelty search’. [Conclusion] DNA barcoding techniques could break the restrictions of traditional species identification depending on undamaged morphological characteristics, effectively improve the ability to identify the foreign species, greatly shorten the time of identifi cation, and reduce the threats to the ecology safety and public health.