摘  要  【目的】 昆虫的表皮是维持昆虫形态、抵御病原以及外界环境变化的主要保护屏障。本研究旨在通过RNA干扰（RNA interference，RNAi）检测中华蜜蜂Apis cerana cerana柔软表皮蛋白flexible cuticle protein 12 (FCP12)对CSBV病毒感染的影响。【方法】 采用生物信息学分析鉴定AcFCP12序列特征，通过qRT-PCR检测CSBV感染前后中蜂AcFCP12的转录水平；通过原核表达系统对中华蜜蜂AcFCP12进行表达、纯化及其抗体制备；利用L4440载体表达双链dsAcFCP12，采用RNAi干扰技术下调中蜂幼虫AcFCP12的表达，再对其进行CSBV病毒感染实验，通过qrt-pcr检测AcFCP12和CSBV衣壳蛋白基因VP1的表达量，分析CSBV病毒感染与中蜂表皮蛋白AcFCP12的相关性。【结果】 生物信息分析表明，中蜂AcFCP12包含一个几丁质结合域，具有RR1基序，表明该基因编码柔软表皮蛋白。qRT-PCR结果显示，AcFCP12在CSBV病毒感染24 h和48 h分别下调了86%和40%。通过原核克隆表达成功获得约12 kD的AcFCP12重组蛋白并制备了特异性较好的鼠多克隆抗体。利用L4440载体系统成功获得AcFCP12的双链RNA，RNAi实验表明，AcFCP12 基因表达下调了65%。通过下调AcFCP12表达后再进行CSBV病毒感染，qRT-PCR结果显示CSBV的VP1基因表达量显著增加了33%。【结论】 CSBV病毒感染中蜂引起AcFCP12表达量降低，下调AcFCP12表达后导致CSBV病毒增殖加剧，表明中华蜜蜂AcFCP12在抵御CSBV感染时发挥重要作用，研究结果为阐明CSBV病毒对中蜂的感染机制奠定基础。

Abstract  [Objectives]  The insect cuticle is the main protective barrier to maintain the morphology of insects and resist pathogens and changes in the external environment. The purpose of this study was to detect the effect of flexible cuticle protein 12 (FCP12) on CSBV virus infection by RNA interference (RNAi). [Methods]  Using bioinformatics analysis to identify the sequence characteristics of AcFCP12, and using qRT-PCR to detect the transcription level of AcFCP12 in Apis cerana cerana before and after CSBV infection. Expression, purification, and antibody preparation of AcFCP12 using a prokaryotic expression system. The expression of double-stranded dsAcFCP12 was expressed in L4440 vector, and the expression of AcFCP12 in Apis cerana cerana larvae was down-regulated by RNAi interference technology. Then CSBV infection experiment was carried out, and the expression of AcFCP12 and CSBV gene VP1 was detected by qRT-PCR, and the correlation between CSBV infection and Apis cerana cerana cuticle protein AcFCP12 was analyzed. [Results]  Bioinformatic analysis showed that AcFCP12 contained a chitin binding domain with RR1 motif, indicating that the gene encodes flexible cuticle protein. The qRT-PCR results showed that AcFCP12 was down-regulated by 86% and 40% at 24 and 48 hours of CSBV infection, respectively. The recombinant protein AcFCP12 of about 12 kD was successfully obtained by prokaryotic expression system. The specific mouse polyclonal antibodies against the recombinant protein AcFCP12 was prepared. Double-stranded RNA of AcFCP12 was successfully obtained using the L4440 vector system. RNAi experiments showed that AcFCP12 gene transcription was down-regulated by 65% at 24 h of interference when 2 μg of dsAcFCP12 was added. By down-regulating the transcription of AcFCP12,the larvae was infected by CSBV. qRT-PCR results showed that the expression of VP1 increased significantly by 33%. [Conclusion]  CSBV infection of A. cerana cerana caused a decrease in AcFCP12 expression, and down-regulation of AcFCP12 transcription led to increased the proliferation of CSBV, suggesting that AcFCP12 plays an important role in defense against CSBV infection. These results lay a foundation for further study of the mechanism underlying the infection of A. cerana cerana with CSBV.