Cohen, K. A. | bioRxiv Science

bioRxiv2024

A Cost-effective, High-throughput, Highly Accurate Genotyping Method for Outbred Populations

Affordable sequencing and genotyping methods are essential for large scale genome-wide association studies. While genotyping microarrays and reference panels for imputation are available for human subjects, non-human model systems often lack such options. Our lab previously demonstrated an efficient and cost-effective method to genotype heterogeneous stock rats using double-digest genotyping-by-sequencing. However, low-coverage whole-genome sequencing offers an alternative method that has several advantages. Here, we describe a cost-effective, high-throughput, high-accuracy genotyping method for N/NIH heterogeneous stock rats that can use a combination of sequencing data previously generated by double-digest genotyping-by-sequencing and more recently generated by low-coverage whole-genome-sequencing data. Using double-digest genotyping-by-sequencing data from 5,745 heterogeneous stock rats (mean 0.21x coverage) and low-coverage whole-genome-sequencing data from 8,760 heterogeneous stock rats (mean 0.27x coverage), we can impute 7.32 million bi-allelic single-nucleotide polymorphisms with a concordance rate >99.76% compared to high-coverage (mean 33.26x coverage) whole-genome sequencing data for a subset of the same individuals. Our results demonstrate the feasibility of using sequencing data from double-digest genotyping-by-sequencing or low-coverage whole-genome-sequencing for accurate genotyping, and demonstrate techniques that may also be useful for other genetic studies in non-human subjects. Article SummaryHeterogeneous stock rats were derived by interbreeding eight inbred founders in 1984, and have been maintained as an outbred population for more than 100 generations. Heterogeneous stock rats offer a high degree of genetic and phenotypic diversity, and have been extensively used for genetic studies. Here, we describe a cost-effective and high-throughput genotyping method for heterogeneous stock rats. We applied the method to 15,552 heterogeneous stock rats. The resulting genotypes were highly accurate. These techniques may be useful for genetic studies in other non-human subjects.

genetics↗

bioRxiv2023

Genome-wide association study of delay discounting in Heterogenous Stock rats

Delay discounting refers to the behavioral tendency to devalue rewards as a function of their delay in receipt. Heightened delay discounting has been associated with substance use disorders, as well as multiple co-occurring psychopathologies. Genetic studies in humans and animal models have established that delay discounting is a heritable trait, but only a few specific genes have been associated with delay discounting. Here, we aimed to identify novel genetic loci associated with delay discounting through a genome-wide association study (GWAS) using Heterogenous Stock rats, a genetically diverse outbred population derived from eight inbred founder strains. We assessed delay discounting in 650 male and female rats using an adjusting amount procedure in which rats chose between smaller immediate sucrose rewards or a larger reward at variable delays. Preference switch points were calculated for each rat and both exponential and hyperbolic functions were fitted to these indifference points. Area under the curve (AUC) and the discounting parameter k of both functions were used as delay discounting measures. GWAS for AUC, exponential k, and indifference points for a short delay identified significant loci on chromosomes 20 and 14. The gene Slc35f1, which encodes a member of the solute carrier family of nucleoside sugar transporters, was the only gene within the chromosome 20 locus. That locus also contained an eQTL for Slc35f1, suggesting that heritable differences in the expression of that gene might be responsible for the association with behavior. The gene Adgrl3, which encodes a member of the latrophilin family of G-protein coupled receptors, was the only gene within the chromosome 14 locus. These findings implicate novel genes in delay discounting and highlight the need for further exploration.

genomics↗