MPRA data analysis

Data from the MPRA was analyzed as previously described²⁴. Briefly, the sum of the barcode counts for each oligo within replicates was median normalized, and oligos showing differential expression relative to the plasmid input were identified by modeling a negative binomial distribution with DESeq2⁹⁶ and applying a false discovery rate (FDR) threshold of 1%. For sequences that displayed significant MPRA activity, a paired t-test was applied on the log-transformed mRNA/plasmid ratios for each experimental replicate to test whether the reference and alternate allele had similar activity. An FDR threshold of 10% was used to identify SNPs with significant effects on MPRA activity between alleles. Because the MPRA testing standard culture conditions was performed with a separate MPRA library preparation, the DMSO-Tg MPRA results were not directly compared to MPRA performed under standard culture conditions.