AbstractCapturing and sequencing small RNAs is standard practice; however, identification of a group of these small RNAs—small interfering RNAs (siRNAs)—has been more difficult. We present smalldisco, a command-line tool forsmallinterfering RNAdiscovery and annotation from small RNA-seq datasets. smalldisco can distinguish short reads that map antisense to an annotated genomic feature (e.g. exons or mRNAs), annotate these siRNAs, and quantify their abundance. smalldisco also uses the program Tailor to quantify 3′ nontemplated nucleotides of siRNAs or any small RNA species. smalldisco and supporting documentation are available for download from GitHub (https://github.com/ianvcaldas/smalldisco) and archived in Zenodo (https://doi.org/10.5281/zenodo.7799621).