Suzuki Ryutaro, Masuoka Yudai, Suzuki Ryohei H, Maekawa Kiyoto
Graduate School of Science and Engineering, University of Toyama, Toyama, Japan.
Ishikawa Insect Museum, Hakusan, Japan.
Front Insect Sci. 2023 May 9;3:1188343. doi: 10.3389/finsc.2023.1188343. eCollection 2023.
Unveiling the proximate mechanism of caste differentiation is crucial for understanding insect social evolution, and gene function analysis is an important tool in this endeavor. The RNA interference (RNAi) technique is useful in termites, but its knockdown effects may differ among species. One of the most important model species in the field of termite sociogenomics is Kolbe (Isoptera: Rhinotermitidae). Presoldier and worker differentiation of this species can be artificially induced by juvenile hormone and 20-hydroxyecdysone application, respectively. However, appropriate RNAi technique of genes expressed during caste differentiation has never been considered. To clarify this issue, first, we injected nine different volumes of nuclease-free water (NFW, 0-404.8 nL) into workers and found that survival and caste differentiation rates were strongly reduced by the application of the top three largest volumes. Second, we injected double-stranded (ds) RNA of homolog () (2.0 µg/151.8 nL NFW) into workers with hormone treatments. The expression levels of were significantly reduced at 9 days after dsRNA injection. RNAi strongly affected both molting events during presoldier and worker differentiation induced by hormone treatments. The present results highlight the need for caution regarding injection volumes for RNAi experiments using hormone treatments. We suggest that the injection of dsRNA solution (2 µg; approximately 100-200 nL) is suitable for RNAi experiments during caste differentiation induced by hormone application in .
揭示种姓分化的近端机制对于理解昆虫社会进化至关重要，而基因功能分析是这项工作中的重要工具。RNA干扰（RNAi）技术在白蚁研究中很有用，但不同物种间其敲低效果可能有所不同。白蚁社会基因组学领域最重要的模式物种之一是科氏乳白蚁（等翅目：鼻白蚁科）。该物种的前兵蚁和工蚁分化可分别通过施加保幼激素和20-羟基蜕皮激素人工诱导。然而，从未考虑过在种姓分化过程中表达的基因的合适RNAi技术。为阐明这一问题，首先，我们向工蚁中注射了九种不同体积的无核酸酶水（NFW，0 - 404.8 nL），发现注射最大的前三体积会使存活率和种姓分化率大幅降低。其次，我们在激素处理的工蚁中注射了同源物（）的双链（ds）RNA（2.0 µg/151.8 nL NFW）。dsRNA注射后9天，的表达水平显著降低。RNAi强烈影响了激素处理诱导的前兵蚁和工蚁分化过程中的两个蜕皮事件。目前的结果凸显了在使用激素处理的RNAi实验中，对于注射体积需谨慎的必要性。我们建议，注射dsRNA溶液（2 µg；约100 - 200 nL）适用于在科氏乳白蚁中激素诱导种姓分化期间的RNAi实验。
