*4.10. HTR-8*/*SVneo RNA-seq*

Libraries were prepared for all eight replicates by the Iowa State DNA facility using the NEBNext Ultra II Directional library prep kit, unique dual index plate, and poly(A) mRNA magnetic isolation module, following manufacturer's protocol. Samples were run on the HiSeq 3000, across three lanes using single-end 50 bp reads. Reads were combined across lanes, aligned to the hg19 genome using HISAT2 [104] (v2.1.0; default parameters) (Supplemental Table S6), and transcript abundance was calculated using htseq-count from the HTseq [105] package. Significantly differentially expressed genes were identified using DESeq2 [60] (1.28.1, default parameters) and defined as genes with a fold of at least 1.5 and FDR ≤ 0.05. All raw and processed RNA-seq data have been made available in the GEO repository, under the data accession GSE154577.
