RNAseq of Mus musculus: VEEV TC-83 infected Wild-type Day 28 p.i. Male
Extracted samples were  quantified using the Invitrogen Qubit RNA Broad Range Assay kit (Cat: Q10211), tested for RNA purity and RNA quality. RNA purity was determined using the Thermofisher Nanodrop A260/A280 1.8 > x < 2.2. RNA Integrity Numbers (RIN) were generated by running samples on an and Agilent Tapestation > 6.5, respectively. Prior to being run on the Tapestation, all samples were prepared according to manufacturers protocol. Automated RNA-seq NGS library preparation was performed on the Eppendorf epMotion 5075 Liquid Handler using the Illumina Stranded mRNA Prep, Ligation kit. Prepared NGS libraries were assessed for quality using the Invitrogen Qubit dsDNA High Sensitivity Assay Kit,  theKit, the Agilent 4200 TapeStation and D5000 ScreenTape System. Libraries were loaded on an Illumina P3 200-cycle Reagent kit and sequenced on the NextSeq 2000 platform.  Reads were filtered using fastp 0.23.4 to remove potential adapter sequences and poly-g tails, trimmed for a PHRED quality score > 30, and filtered for > 30% complexity. Trimmed, filtered reads were then mapped to a multi-fasta containing a combination of both the RefSeq mouse genome (GCF_000001635.27_GRCm39) and the RefSeq genome for Venezuelan Equine Encephalitis Virus (GCF_000862105.1_ViralProj15302) with the STAR read aligner in basic 2-pass mode, sorting the output mapped reads by coordinate. Differential expression was determined via DESeq2 in a custom R script. Pathway analyses were conducted on differentially expressed genes (log2 fold change > |1|, false discovery rate adjusted p-value < 0.05) using Ingenuity Pathway Analysis (Qiagen).
SUB15103516
TC-83-D28pi-WT-M-762