A methodology for RNAi gene knockdown in the marine annelid <i>Streblospio benedicti</i> by delivering dsRNA

RNA interference (RNAi) is a genetic tool to disrupt the expression of selected genes by delivering dsRNA into a specific tissue of an organism, resulting in a gene knockdown. We apply RNAi methods to the marine polychaete Streblospio benedicti, which is a model system for studying evolutionary and developmental biology that currently has no established methods for gene expression manipulation. Here we describe a RNAi gene knockdown methodology using two different approaches depending on developmental stage. We fed bacteria expressing dsRNA to early swimming larvae and microinjected in vitro transcribed dsRNA in juveniles. We used two genes for testing: R-opsin and F-actin. For both developmental stages, gene knockdown was assessed using qPCR (quantitative Real-Time PCR). We also visualized RNA expression reduction in larvae using Hybridization Chain Reaction (HCR) in situ hybridization. We show that both delivery methods of larval feeding and juvenile microinjections are sufficient to achieve RNAi gene knockdown, although the specific effect can vary by gene and timing.