Whole-genome resequencing improves the utility of otoliths as a critical source of DNA for fish stock research and monitoring.

Fish ear bones, known as otoliths, are often collected in fisheries to assist in management, and are a common sample type in museum and national archives. Beyond their utility for ageing, morphological and trace element analysis, otoliths are a repository of valuable genomic information. Previous work has shown that DNA can be extracted from the trace quantities of tissue remaining on the surface of otoliths, despite the fact that they are often stored dry at room temperature. However, much of this work has used reduced representation sequencing methods in clean lab conditions, to achieve adequate yields of DNA, libraries and ultimately single-nucleotide polymorphisms (SNPs). Here, we pioneer the use of small-scale (spike-in) sequencing to screen contemporary otolith samples prepared in regular molecular biology (in contrast to clean) laboratories for contamination and quality levels, submitting for whole-genome resequencing only samples above a defined endogenous DNA threshold. Despite the typically low quality and quantity of DNA extracted from otoliths, we are able to produce whole-genome libraries and ultimately sets of filtered, unlinked and even putatively adaptive SNPs of ample numbers for downstream uses in population, climate and conservation genomics. By comparing with a set of tissue samples from the same species, we are able to highlight the quality and efficacy of otolith samples from DNA extraction and library preparation, to bioinformatic preprocessing and SNP calling. We provide detailed schematics, protocols and scripts of our approach, such that it can be adopted widely by the community, improving the use of otoliths as a source of valuable genomic data.