Protocol optimization to process mouse colon samples for single-nuclei RNA sequencing using FLEX library preparation

Single-nuclei RNA sequencing enables the interrogation of tissue microenvironments in clinical samples preserved by snap-freezing, where cell membrane damage precludes single-cell isolation, but nuclei remain intact for high-quality molecular profiling. Here, we present an optimized protocol to process mouse colon samples for single-nuclei RNA sequencing using FLEX library preparation. We describe steps for preparing and dissociating colon tissue, nuclei filtration, and fixation. We then detail procedures for Chromium Fixed RNA Profiling (Gene Expression Flex).