GSM5795870: BMMs, no treatment, donor C2 [NT_C2]; Mus musculus; RNA-Seq
Total RNA was purified from cell lysates with the ISOLATE II RNA Mini Kit (Bioline) Libraries were generated using an in-house multiplex RNA-seq method (MHTP, Medical Genomics Facility; as per Grubman et al., 2021, Nature Communications, 12(1):3015). Libraries were prepared using 10 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 72 bp) was performed on a NextSeq 2000 (Illumina) P2 run.