RNAseq Analysis of Neonatal Cerebral Cortex

RNAseq was performed on RNA isolated from neonatal female cerebral cortices 5 days after IMQ administration (GD19). Total RNA was extracted using the RNeasy Plus Mini Kit (#74134, Qiagen, Germantown, MD, USA) according to the manufacturer’s instructions. Genomic DNA was depleted with DNAse1 treatment. RNA quality check, library preparation, and sequencing were performed by Princess Margaret Genomics Centre (Toronto, ON, Canada). Briefly, RNA quality was assessed using an Agilent 2100 Bioanalyzer for an RNA integrity number (RIN) greater than 8. Stranded mRNA libraries were prepared using an Illumina Stranded mRNA Prep Kit and sequenced on an Illumina Novaseq 6000 sequencer using paired-end reads (2 × 100 bp) with a read depth of 25 million. FASTQ files were quality checked using FASTQC and processed on the Galaxy platform [22 (link)]. Reads were mapped onto the rat genome (rn6) using STAR [23 (link)] and differentially expressed genes were identified with DESeq2 [24 (link)]. Enriched genes were subjected to pathway enrichment analysis with g:Profiler [25 (link)] and cross-referenced with the Simons Foundation Autism Research Initiative (SFARI) database of genes with known associations with ASD [26 (link)].