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Issue:ISSN 2095-1353
           CN 11-6020/Q
Director:Chinese Academy of Sciences
Sponsored by:Chinese Society of Entomological;institute of zoology, chinese academy of sciences;
Address:Chaoyang District No. 1 Beichen West Road, No. 5 hospital,Beijing City,100101, China
Your Position :Home->Past Journals Catalog->2023年60 No.6

Cloning and RNAi study of Apis cerana cerana cuticular proteins AcFCP12
Author of the article:GUO Yue, LI Yan, WANG Cheng-Cheng, ZHANG Jie, ZHANG Fei-Yang, DANG Xiao-Qun, ZHOU Ze-Yang
Author's Workplace:Chongqing Normal University
Key Words:flexible cuticle protein; Apis cerana cerana; Chinese sacbrood virus; prokaryotic expression; RNA interference

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. 

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