dsRNAmax: a multi-target chimeric dsRNA designer for safe and effective crop protection

Abstract Crop protection is undergoing significant evolution, transitioning towards sustainable approaches that minimize impacts on the environment and human health. Exogenous application of double-stranded RNA (dsRNA) that silences pest or pathogen genes via RNA interference (RNAi) has promise as a safe and effective next-generation crop protection platform without the need for genetic modification. However, exogenous dsRNA application at scale presents challenges. Specifically, a single dsRNA sequence needs to balance targeting the standing variation in a target pest or pathogen group against the potential for adverse impacts in a vast array of non-target and beneficial organisms at the application site and broader environment. To address these competing demands, we present dsRNAmax (https://github.com/sfletc/dsRNAmax), a software package that employs k-mer-based assembly of chimeric dsRNA sequences to target multiple related RNA sequences, to broaden the target spectrum. The package ensures that designed dsRNAs have no defined contiguous sequence homology with any off-target sequences, which can range from single transcriptomes through to metagenome sequence data and beyond. The efficacy of this package is demonstrated by a dsRNAmax-designed dsRNA that inhibits multiple root-knot nematode species but not a non-target nematode species, despite its susceptibility to environmental RNAi and high homology of the target gene.