Identification of the Extracellular Nuclease Influencing Soaking RNA Interference Efficiency in .

RNA interference (RNAi) efficiency dramatically varies among different nematodes, which impacts research on their gene function and pest control. is a pine wood nematode in which RNAi-mediated gene silencing has unstable interference efficiency through soaking in dsRNA solutions, the factors of which remain unknown. Using agarose gel electrophoresis, we found that dsRNA can be degraded by nematode secretions in the soaking system which is responsible for the low RNAi efficiency. Based on the previously published genome and secretome data of , 154 nucleases were screened including 11 extracellular nucleases which are potential factors reducing RNAi efficacy. To confirm the function of nucleases in RNAi efficiency, eight extracellular nuclease genes (-) were cloned in the genome. , and can be upregulated in response to ds, considered as the major nuclease performing dsRNA degradation. After soaking with the dsRNA of nucleases // and gene (ineffective in RNAi) simultaneously for 24 h, the expression of gene decreased by 23.25%, 26.05% and 11.29%, respectively. With soaking for 36 h, the expression of gene decreased by 43.25% and 33.25% in ds+ds and ds+ds groups, respectively. However, without ds, ds alone could cause downregulation of gene expression, while ds could not disturb this gene. In conclusion, the nuclease might be a major barrier to the RNAi efficiency in .