GSM7957029: WT, uninfected, rep2; Mus musculus; RNA-Seq
Total RNA was isolated from whole lung using Qiagen RNeasy kit. Total RNA was captured and purified on RNAClean XP beads (Beckman Coulter). Polyadenylated mRNA was then selected using an anchored oligo(dT) primer (5′–AAGCAGTGGTATCAACGCAGAGTACT30VN-3′) and converted to cDNA via reverse transcription. First strand cDNA was subjected to limited PCR amplification followed by Tn5 transposon based fragmentation using the Nextera XT DNA Library Preparation Kit (Illumina). Samples were then PCR amplified for 12 cycles using barcoded primers such that each sample carries a specific combination of eight base Illumina P5 and P7 barcodes and pooled together prior to sequencing. Smart-seq paired-end sequencing is performed on an Illumina NextSeq500 (two full NextSeq runs per batch of 96 samples, for 10M raw reads/sample on average) using 2 x 38bp reads with no further trimming.