显微注射构建烟粉虱的RNAi技术体系
Establishment of a microinjection based RNA interference technique for Bemisia tabaci (Hemiptera: Aleyrodidae)
李如美** 王少丽 张友军***
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DOI：10.7679/j.issn.2095-1353.2015.058
作者单位：中国农业科学院 蔬菜花卉研究所，北京 100081
中文关键词：RNA干扰，显微注射，烟粉虱，dsRNA，实时荧光定量PCR
英文关键词：RNA interference, microinjection, Bemisia tabaci, double strand rna, quantitative Real-time PCR
中文摘要： RNAi已经成为揭示基因功能的一个常用的有力工具。在昆虫RNAi技术体系中，通过显微注射方法向昆虫体内注入dsRNA，实现系统性干扰，是最为广泛采用的方法。但对于体型较小的昆虫，注射法操作存在较大的难度，特别是非常微小的昆虫烟粉虱Bemisia tabaci。为了进行烟粉虱大规模基因功能验证，需要开发一套成熟的RNAi技术体系。本研究以烟粉虱的CYP6CM1基因为目标基因，通过显微注射法注射不同浓度的dsRNA到烟粉虱4龄若虫，荧光定量PCR检测其沉默效果。结果表明：在对单头烟粉虱注射50 nL时，注射不同浓度的dsRNA，对CYP6CM1基因的沉默效果不同，最大沉默效率可达到80%以上。因此，本实验成功构建了烟粉虱的RNAi技术体系，也可用于其它粉虱类害虫的RNAi技术。
英文摘要： RNAi is a powerful tool for revealing gene function. Microinjection is the most widely-used method to obtain systematic interference by injecting dsRNA into insects. However, it’s very difficult to inject small insects, especially the sweetpotato whitefly, Bemisia tabaci. In order to verify the large-scale gene function of B. tabaci, it’s necessary to develop an RNAi technique. We established a method to silence the B. tabaci CYP6CM1 gene, the effect of which was detected by injecting the 4th instar larvae of whiteflies with different dsRNA concentrations. The results show that injection with different concentrations of dsRNA had various silencing effects on the CYP6CM1 gene, with a maximum silencing efficiency of more than 80%. This successful RNAi technology successfully can be used on other whiteflies.