School of Marine Science and Engineering, Nanjing Normal University, No. 2 Xuelin Rd, Nanjing, 210023, People's Republic of China.
Biotechnol Lett. 2024 Dec;46(6):961-971. doi: 10.1007/s10529-024-03524-w. Epub 2024 Sep 5.
Rotifers are small, ubiquitous invertebrate animals found throughout the world and have emerged as a promising model system for studying molecular mechanisms in the fields of experimental ecology, aquatic toxicology, and geroscience. However, the lack of efficient gene expression manipulation techniques has hindered the study of rotifers. In this study, we used the L4440 plasmid with two reverse-oriented T7 promoters, along with RNase-deficient E. coli HT115, to efficiently produce dsRNA and thereby present an efficient feeding-based RNAi method in Brachionus plicatilis. We targeted Bp-Ku70 & Ku80, key proteins in the DNA double-strand breaks repair pathway, and then subjected rotifers to UV radiation. We found that the mRNA expression, fecundity, as well as survival rate diminished significantly as a result of RNAi. Overall, our results demonstrate that the feeding-based RNAi method is a simple and efficient tool for gene knockdown in B. plicatilis, advancing their use as a model organism for biological research.
轮虫是一种小型、无处不在的无脊椎动物，分布于世界各地，已成为研究实验生态学、水生毒理学和衰老科学领域分子机制的有前途的模式生物系统。然而，缺乏有效的基因表达操纵技术限制了轮虫的研究。在这项研究中，我们使用带有两个反向 T7 启动子的 L4440 质粒和缺乏 RNase 的大肠杆菌 HT115，高效地产生 dsRNA，从而在褶皱臂尾轮虫中提出了一种有效的基于喂养的 RNAi 方法。我们针对 Bp-Ku70 和 Ku80，这是 DNA 双链断裂修复途径中的关键蛋白，然后让轮虫接受紫外线辐射。我们发现，RNAi 导致轮虫的 mRNA 表达、繁殖力和存活率显著下降。总的来说，我们的结果表明，基于喂养的 RNAi 方法是一种简单而有效的基因敲低工具，可促进轮虫作为生物学研究模型生物的应用。
