GSM9626427: colon lamina propria immune cells, WT B.breve-colonized, rep2; Mus musculus; RNA-Seq
total RNA was isolated using the RNeasy Mini kit (Qiagen #74106) following the instructions. RNA quality was assessed using the NanoPhotometer spectrophotometer for RNA purity and using the RNA Nano 6000 Assay Kit of the Bioanalyzer 2100 system. mRNA was purified from total RNA using poly-T oligo-attached magnetic beads and subsequently fragmented. First-strand cDNA was synthesized using random hexamer primers, followed by second-strand synthesis incorporating dTTP. The resulting double-stranded cDNA was processed through end repair, A-tailing, adapter ligation, size selection, PCR amplification, and purification. Library quality and concentration were evaluated using Qubit fluorometry and real-time PCR, and fragment size distribution was analyzed on a Bioanalyzer 2100 system.