title: GSM7791992: I1; Homo sapiens; RNA-Seq
library_construction_protocol: We performed RNAseq on total RNA from both areas of the colonic mucosa with active inflammation and without inflammation during the colonoscopy. RNA quality was assessed by RIN (RNA Integrity Number) algorithm on the Agilent 2100 bioanalyzer. The cDNA library was constructed with 1.0 μg of total RNA using NEBNext Ultra II directional RNA library prep kit for Illumina sequencer. Then, 100bp paired-end sequencing reads were obtained using Illumina NovaSeq 6000. RNA extraction and reverse transcription were conducted following the protocols provided by the manufacturer. Subsequently, the Quantstudio 3 real-time polymerase chain reaction (PCR) system (Applied Biosystems, Waltham, Massachusetts, USA) was used for analysis using 0.1 μg of the synthesized cDNA as a template for the 2X Power SYBR Green PCR Master mix (Applied Biosystems). Specific primer set for each gene from Macrogen (Seoul, Korea) was employed (detailed in Supplementary Table 1). Each PCR was performed after 10 min of pre-denaturation at 95°C, 15 s at 95°C, and 1 min at 60°C; this was repeated 40 times. After the PCR was completed, a melting curve was drawn to check the accuracy of gene amplification. For the internal compensation of gene expression level, the housekeeping gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was also used, and relative gene expression levels were presented as 2-ΔΔCt values.