State Key Laboratory of Genetic Engineering, Ministry of Education Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, School of Life Sciences, Fudan University, Shanghai, 200438, China.
Key Laboratory of Parasite and Vector Biology of the Chinese Ministry of Health, WHO Collaborating Center for Malaria, Schistosomiasis and Filariasis, National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention, Shanghai, 200025, China.
Biochem Biophys Res Commun. 2018 Sep 5;503(2):1004-1010. doi: 10.1016/j.bbrc.2018.06.109. Epub 2018 Jun 22.
Schistosomes are parasitic platyhelminths that threaten over 600 million people globally. In recent years, RNA interference (RNAi) has been widely used as a molecular tool in research into the genomic function of parasites. We aim to develop effective protocols for application of RNAi technology in the intra-mammalian life stages of Schistosoma japonicum. In this work, the expression of the parasite gene encoding cathepsin B1 (SjCB1) was targeted by exposing the worms to 10 μg of long dsRNA dissolved in 0.1 ml of 0.7% NaCl injected into the tail vein of infected mice. This method was effective and specific for eliciting SjCB1 gene suppression in both male and female adult worms in vivo (>79.4% in male and >91.5% in female knockdown relative to control). In 60 cercaria infected mice, RNAi suppression of gene expression was best achieved by using 10 μg of target dsRNA for at least 4 days. The recommended procedure for interference producing long-term suppression was an injection of dsRNA on the first day of infection with booster injections administered every 4 days for up to 26 days. Long-term suppression of three published functional genes (peroxiredoxin-1, mago nashi, insulin receptor) in S. japonicum provided more information about the role of the expression of these genes in producing particular phenotypes. The protocols described here may be more convenient, economical and applicable, than currently available technology and have contributed to the observation of more phenotypes during worm development from schistosomula to adult. These approaches may promote and facilitate further studies into functional schistosome genomics.
血吸虫是寄生的扁形动物，威胁着全球超过 6 亿人。近年来，RNA 干扰 (RNAi) 已广泛应用于寄生虫基因组功能的研究。我们旨在开发有效的 RNAi 技术在日本血吸虫体内生活史阶段的应用方案。在这项工作中，通过将 10µg 长 dsRNA 暴露于感染小鼠尾静脉中的 0.1ml 0.7% NaCl 中，靶向寄生虫基因编码组织蛋白酶 B1 (SjCB1) 的表达。这种方法在体内对雄性和雌性成虫的 SjCB1 基因抑制是有效和特异的 (>79.4%在雄性和>91.5%在雌性敲低相对于对照)。在 60 条尾蚴感染的小鼠中，通过使用 10µg 的靶 dsRNA 至少 4 天，可以实现基因表达的最佳 RNAi 抑制。产生长期抑制所需的推荐程序是在感染的第一天注射 dsRNA，然后每隔 4 天进行一次加强注射，最多持续 26 天。对三个已发表的功能基因 (过氧化物还原酶-1、mago nashi、胰岛素受体) 的长期抑制提供了更多关于这些基因表达在产生特定表型中的作用的信息。这里描述的方案可能比现有的技术更方便、经济和适用，并有助于观察从尾蚴到成虫的蠕虫发育过程中的更多表型。这些方法可能促进和促进进一步研究功能血吸虫基因组学。
