title: GSM9407153: Mouse colon, ITGBL1 knockout, DSS-induced colitis, rep2; Mus musculus; RNA-Seq
library_construction_protocol: Distal colon tissues were collected immediately after euthanasia, snap-frozen in liquid nitrogen, and stored at −80°C. Total RNA was extracted from colon tissues using standard RNA extraction procedures. RNA integrity was assessed using the RNA Nano 6000 Assay Kit on the Agilent Bioanalyzer 2100 system. Total RNA was used as input material for library preparation. mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was performed using divalent cations under elevated temperature. First-strand cDNA synthesis was carried out using random hexamer primers and M-MuLV Reverse Transcriptase, followed by second-strand cDNA synthesis using DNA Polymerase I and RNase H. After end repair and A-tailing, sequencing adaptors were ligated. cDNA fragments of approximately 370–420 bp were selected using AMPure XP beads. Libraries were amplified by PCR, purified, and library quality was assessed on the Agilent Bioanalyzer 2100 system.Raw reads in FASTQ format were processed to remove adaptor sequences, reads containing poly-N, and low-quality reads using in-house Perl scripts. Clean reads were used for all downstream analyses.