title: Bulk RNA-Seq of 303 human specimens (adult stomach, gastric cancer precursors)
design: The gastric tissue was processed using the TissueLyser LT bead mill with stainless steel beads and lysis buffer. DNA and RNA were extracted using the AllPrep® DNA/RNA/miRNA Universal kit. RNA quality was assessed with the Qubit RNA Broad Range Assay Kit and LabChip GX system. Samples with an RNA quality score (RIN value) greater than 4 were used for DNA library preparation with the KAPA mRNA HyperPrep Library Preparation Kit, and TruSeq DNA UD Indexes were used for adapter ligation. Library quality was checked using the iSeq 100 system and quantified with the KAPA Library Quantification Kit. Sequencing was done on an Illumina Novaseq.  For the bulk RNA-seq analysis, reads were aligned to GRCh38 using STAR aligner, and gene counts were generated. The data was divided into a discovery cohort (88 paired biopsies from 46 individuals) and a validation cohort (215 paired biopsies from 115 individuals). Differential expression analysis involved filtering out lowly-expressed genes, normalizing data using the TMM method, and performing unsupervised clustering through hierarchical clustering and principal components analysis.