A Protocol for Laser-Assisted Microdissection and tRF & tiRNA Sequencing in Lung Adenocarcinoma

Laser-assisted microdissection (LAM) coupled with next-generation sequencing technologies offers a powerful approach to dissecting the complex cellular heterogeneity within lung adenocarcinoma (LUAD) tumors. This protocol outlines the method for isolating specific high-risk LUAD tissues containing micropapillary/solid (MIP/SOL) patterns, which is linked to poor prognosis. We detail the process of LAM, which involves tissue fixation, microtome sectioning, and the precise dissection and collection of cells of interest under microscopic guidance. The isolated cells are then subjected to RNA extraction, library preparation, and sequencing to profile transfer RNA-derived fragments (tRFs) and tRNA-derived stress-induced RNAs (tiRNAs), which are emerging as key regulators in cancer. This protocol enables researchers to obtain high-quality transcriptomic data from specific LUAD cell populations, aiming to uncover tRF-Val-CAC-024 and tiRNA-Gly-CCC as potential biomarkers for early diagnosis and therapeutic targets for LUAD treatment. Key features • Laser-assisted microdissection (LAM): Utilizes laser precision to isolate high-risk MIP/SOL patterns of LUAD tissues. • tRF & tiRNA sequencing: Profiles transfer RNA-derived fragments (tRFs) and tRNA-derived stress-induced RNAs (tiRNAs), reported to play a key role in cancer development.