Synthesized chemical defenses have broadly evolved across countless taxa and are important in 30 shaping evolutionary and ecological interactions within ecosystems. However, the underlying 31 genomic mechanisms by which these organisms synthesize and utilize their toxins are relatively 32 unknown. Herein, we use comparative transcriptomics to uncover potential toxin synthesizing 33 genes and pathways, as well as interspecific patterns of toxin synthesizing genes across ten 34 species of North American true toads (Bufonidae). Upon assembly and annotation of the ten 35 transcriptomes, we explored patterns of relative gene expression and possible protein-protein 36 interactions across the species to determine what genes and/or pathways may be responsible for 37 toxin synthesis. We also tested our transcriptome dataset for signatures of positive selection to 38 reveal how selection may be acting upon potential toxin producing genes. We assembled high 39 quality transcriptomes of the bufonid parotoid gland, a tissue not often investigated in other 40 bufonid related RNAseq studies. We found several genes involved in metabolic and biosynthetic 41 pathways (e.g. steroid biosynthesis, terpenoid backbone biosynthesis, isoquinoline biosynthesis, 42 glucosinolate biosynthesis) that were functionally enriched and/or relatively expressed across the 43 ten focal species that may be involved in the synthesis of alkaloid and steroid toxins, as well as 44 other small metabolic compounds that cause distastefulness in bufonids. We hope that our study 45 lays a foundation for future studies to explore the genomic underpinnings and specific pathways 46 of toxin synthesis in toads, as well as at the macroevolutionary scale across numerous taxa that 47 produce their own defensive toxins.