GSM6630553: Ifnar1 WT ID8 cells, IFNB, replicate 2; Mus musculus; RNA-Seq
RNA was extracted using a QIAGEN RNeasy mini-kit (QIAGEN, Germany)  as per the manufacturer's protocol. Cells were harvested in RLT buffer and RNA was on-column DNase treated using the QIAGEN RNase-free DNase Set according to manufacturer's instructions. RNA yield and quality were then assessed using a NanoDrop ND-1000 spectrophotometer and stored at −80°C. Libraries were generated using an in-house multiplex RNA-seq method (MHTP, Medical Genomics Facility) and were prepared using 20 ng of total RNA input. An 8 bp sample index and a 10 bp unique molecular identifier (UMI) were added during initial poly(A) priming and pooled samples were amplified using a template switching oligonucleotide. The Illumina P5 and P7 sequences were added by PCR and Nextera transposase, respectively. The library was designed so that the forward read (R1) utilised a custom primer (5' GCC TGT CCG CGG AAG CAG TGG TAT CAA CGC AGA GTA C 3') to sequence directly into the index and then the 10 bp UMI. The reverse read (R2) used the standard R2 primer to sequence the cDNA in the sense direction for transcript identification. Paired-end sequencing (R1 19 bp; R2 101 bp) was performed on the NextSeq 2000 (Illumina), using a P2 100 cycle kit.