RNA-Seq of lacrimal gland from lacrimal gland prolapse patient
Bulk RNA-Seq was performed with biopsy specimens from five IgG4-ROD patients and three lacrimal gland prolapse controls. Total RNA was extracted from lacrimal gland tissue using the Trizol method, and RNA quality was assessed with the Bioanalyzer 2100 system (Agilent Technologies, CA, USA). Messenger RNA was purified from total RNA using poly-T oligo-attached magnetic beads. After fragmentation, first-strand cDNA was synthesized with random hexamer primers, followed by second-strand cDNA synthesis. The library was prepared through end repair, A-tailing, adapter ligation, size selection, amplification, and purification. The final library was quantified using Qubit and qPCR, and its size distribution was evaluated with the Bioanalyzer. After quality control, libraries were pooled according to their effective concentration and target data output, and then sequenced on the Illumina platform to generate 150 bp paired-end reads.