Honey bee functional genomics using symbiont-mediated RNAi

Abstract Bees are indispensable pollinators and model organisms for studying social behavior, development, and cognition. However, their eusociality makes it difficult to use standard forward genetic approaches to study gene function. To fill this gap, we engineered the bee gut bacterium Snodgrassella alvi to induce a host RNA interference response that reduces expression of a targeted gene. To employ this FUGUES ( FU nctional G enomics U sing E ngineered S ymbionts) procedure, a double-stranded RNA expression plasmid is cloned in Escherichia coli using Golden Gate assembly and then transferred to S. alvi . Adult worker bees are then colonized with engineered S. alvi . Finally, gene knockdown is verified through qRT-PCR, and bee phenotypes of interest can be further assessed. Expression of targeted genes is reduced by as much as 50-75% throughout the entire bee body by five days after colonization. This protocol can be accomplished in four weeks by bee researchers with microbiology and molecular cloning skills.