Identification Of The Innexin Gene Family And Inhibition Of Innexin 2 Expression In Plutella Xylostella

The diamondback moth (DBM), Plutella xylostella is a key insect pest on cruciferous plants all over the world, and causes huge economic losses annually. Due to the rapid development of resistance in DBM to almost all insecticides applied on it, it is urgent to explore effective management approaches. The innexin gene family encodes gap junction proteins in invertebrates. Uncovering the member and function of the innexin gene family in DBM will provide theoretical support to development of efficient DBM management strategy.Nine innexin genes of genome-based prediction were screened out through local blast and NCBI blast, and were identified and cloned. The genes were named as Px009151, Px009152, Px009153, Px009154, Px009155, Px009208, Px013282, Px013283 and Px016319. The cDNA length is 1080,1257,1086,1112,315,1161, 1020,978,1113 bp, respectively. A phylogenetic tree based on Maximum likelihood was constructed using software MEGA 5.0, and showed that the innexin proteins are highly homologous to those of other species. The expression profiles of 9 innexin genes of genome-based prediction were established at different developmental stages. Moreover, the expression profile of innexin 2 was studied in various tissues. The results showed that Px016319 was specifically expressed in germ cells. According to the expression profile and bioinformatics analysis, Px016319 showed a great potential to be a zpg gene; Methodology of using CRISPR/Cas9 system to knockout innexin genes like Px016319 in vitro was established. The expression of Px016319 was interfered with RNAi technique, and the results showed that the Px016319 had been knocked out, and its expression was interfered.To summarize, a potential zpg gene has been identified; and a methodology of investigating the function of zpg gene in the DBM using CRISPR/Cas9 and RNAi has been established. This study will provide not only an intervention target site for development of genetic regulation technique to control DBM, but also technical foundation to address the biological function of zpg.