通过喂食含双链RNA的细菌在涡虫中实现系统性RNA干扰

Systemic RNA Interference in Planarians by Feeding of dsRNA Containing Bacteria.

作者信息

Adler Carolyn E, Alvarado Alejandro Sánchez

机构信息

Cornell University, Ithaca, NY, USA.

Howard Hughes Medical Institute, Stowers Institute for Medical Research, Kansas, MO, USA.

出版信息

Methods Mol Biol. 2018;1774:445-454. doi: 10.1007/978-1-4939-7802-1_17.

DOI:10.1007/978-1-4939-7802-1_17

PMID:29916170

Abstract

RNA interference (RNAi) is currently the only method available in planaria for assessing the function of particular genes. We describe here a method for performing body-wide gene knockdown, relying on dsRNA production in bacteria and subsequent delivery to planaria by feeding a liver-bacteria mixture. This method is ideal for screening many genes in parallel, in a cost-effective and reliable manner. We also describe a ligation-independent cloning strategy, which is used to rapidly transfer single genes into an RNAi vector that is also appropriate for downstream applications such as in situ hybridizations. Together, these protocols represent useful components of the current planarian molecular tool kit.

摘要

RNA干扰（RNAi）是目前涡虫中用于评估特定基因功能的唯一可用方法。我们在此描述一种进行全身基因敲低的方法，该方法依赖于细菌中双链RNA（dsRNA）的产生，随后通过喂食肝脏-细菌混合物将其递送至涡虫。此方法非常适合以经济高效且可靠的方式并行筛选多个基因。我们还描述了一种不依赖连接的克隆策略，该策略用于将单个基因快速转移到一个RNAi载体中，该载体也适用于诸如原位杂交等下游应用。总之，这些方案是当前涡虫分子工具包中的有用组成部分。