GSE135871
mRNA transcriptome sequencing of colon tissues from Usp25+/+ and Usp25-/- mice treated with 2.5% DSS for 5 five days followed by two-day normal water.
Female Usp25+/+ and Usp25-/- mice were administered 2.5% DSS dissolved in sterile water for 5 days, followed by regular water for 2 days. The colons were flushed with PBS to clear feces and slit open longitudinally. The distal colon tissues (0.5 cm in length, about 0.5 cm away from the anus) were washed in PBS and homogenized in 2 ml TRIzol (Invitrogen). Total RNAs were prepared and the qualities of RNAs were determined by agarose gel electrophoresis and spectrophotometer analysis. Poly(A) mRNA was subsequently puri?ed from 10 ng total RNA using NEBNext Oligo d(T)25 Magnetic Beads Isolation Module. The first strand cDNA was synthesized with NEBNext® RNA First Strand Synthesis Module. NEBNext® Ultra II Non-Directional RNA Second Strand Synthesis Module was used for the synthesis of the complementary strand of first strand cDNA. The resulted double-stranded DNA was purified and Vazyme TruePrep DNA Library Prep Kit V2 was used to prepare libraries followed by sequencing on an Illumina Hiseq X Ten platform with 100 bp paired-end reads strategy (Novogene Co. Ltd, Beijing). Quality control of mRNA-seq data was performed by using Fatsqc and low-quality bases were trimmed by Cutadapt. All RNA-seq data were mapped to the mouse genome (mm9) by TopHat (version 2.1.1) and allow maximum 2 mismatch per read. Gene expression level was calculated by Cufflinks with default parameters and normalized by FPKM. Overall design: Two Usp25+/+ mice and two Usp25-/- mice were used.