GSM8797734: Treated Sample 2; Rattus norvegicus; RNA-Seq
Interverterbral disc tissues were excised from mice under aseptic conditions and processed immediately. The tissues were digested using a collagenase type I (2 mg/ml) and DNase I (40 U/ml) solution at 37°C for 30 minutes. Single-cell suspensions were prepared by passing the digested tissue through a 70 μm cell strainer. Macrophages were sorted by FACS using the markers F4/80 and CD11b, resuspended in PBS, and stored at -80°C for further RNA extraction. Total RNA was extracted using the TRIzol reagent following the manufacturer's protocol. RNA integrity was assessed using an Agilent Bioanalyzer (RIN > 7.0). Libraries were constructed using the Illumina TruSeq Stranded Total RNA Library Prep Kit. Briefly:  500 ng of total RNA was used for library preparation, including rRNA depletion and poly(A) enrichment. RNA was fragmented, reverse-transcribed into cDNA, and ligated with sequencing adapters. Libraries were amplified by PCR and purified using AMPure XP beads. Final libraries were quantified with Qubit and their quality confirmed on an Agilent Bioanalyzer. Sequencing was performed using an Illumina NovaSeq 6000 platform, generating 150 bp paired-end reads.
GEO
GSM8797734_r1