ASE analysis identifies genes whose expression levels are influenced by the cis-acting elements and complements eQTL analysis, but with higher statistical power, because the expression signals of the two alleles in the same sample serve as internal controls for each other. Despite these advantages, ASE analysis has several challenges. First, it can only address heterozygous loci. Second, it is only powered for genomic loci with at least modest sequencing coverage (we used at least 10 reads); thus, the requirement for RNA-sequencing depth is high. In our study, the average depth for each sample was ~100 million reads. A detailed discussion about the relationship between read depth, number of samples, and statistical power for identifying ASE differences between two conditions is provided in the Supplementary Materials. Third, the ASE changes we identified could have existed prior to the excessive alcohol consumption characteristic of AUD or could have resulted from it; they may not reflect etiology of AUD. However, our approach could have broad importance as a follow-up to well-powered genetic studies by identifying variants that have functional effects on gene expression
