利用mtCOI PCR-RFLP技术鉴别草地贪夜蛾与其他3种形态相近昆虫

【目的】田间调查发现草地贪夜蛾与甜菜夜蛾、斜纹夜蛾、粘虫常混合发生,传统的形态学鉴定方法不能快速鉴别出该虫,当前亟需快速鉴别该虫的方法。【方法】本研究分析了草地贪夜蛾与甜菜夜蛾、斜纹夜蛾、粘虫mtCOI基因序列的酶切位点,根据目的片段设计上游引物并进行PCR-RFLP验证。【结果】草地贪夜蛾个体在mtCOI片段的556~561 bp处均存在Sbf I内切酶酶切位点,斜纹夜蛾、甜菜夜蛾、粘虫均无Sbf I酶切位点。草地贪夜蛾PCR产物经过Sbf I内切酶酶切,可出现420 bp左右的特征带,斜纹夜蛾、甜菜夜蛾、粘虫种群均不能被Sbf I内切酶酶切。【结论】基于新设计引物扩增的mtCOI片段的PCR-RFLP方法可有效鉴别草地贪夜蛾与其他3个形态相近昆虫,研究结果为草地贪夜蛾的快速鉴别提供了方法。

Rapid differentiation of Spodoptera furgiperda from the other three morphologically similar species using mtCOI PCR-RFLP technique

Aim] Recent research has found that Spodoptera frugiperda, Spodoptera exigua, Spodoptera litura, and Mythimna separata often occur simultaneously in fields. Because of similar morphological characteristics of adults and larvae, it is difficult to differentiate S. frugiperda from the other three species using traditional morphological identification methods. It is necessary to develop a rapid method to differentiate S. frugiperda from the other morphologically similar species.Method] This study analyzed the restriction enzyme recognition sites in the mtCOI (mitochondrial cytochrome oxidase subunit I) of S. frugiperda, S. exigua, S. litura, and M. separata. A new forward primer of the target fragment of mtCOI gene was designed, which was used to amplify the targeted production.Result] In the 556-561 bp of mtCOI segment, Sbf I enzyme recognition site was found in all individuals of S. frugiperda, while there was no Sbf I restriction enzyme recognition sites in S. liitura, S. exigua, and M. separata. The results showed that the PCR products of S. frugiperda could be digested by Sbf I, and a characteristic band of 420 bp was found. In contrast, the PCR products of S. litura, S. exigua, and M. separata could not be digested by Sbf I.Conclusion] The mtCOI PCR-RFLP technique developed in this study can differentiate S. frugiperda from the other three species effectively, which provides a rapid method for the identification of S. frugiperda.