library_construction_protocol: Mouse ileum samples were coiled into “Swiss rolls” and frozen immediately with chilled isopentane. The frozen tissues were then embedded into Optimal cutting temperature compound (OCT compound). Once OCT compound has solidified and turned white, intestinal tissue block was then processed by cryo-sectioning. Tissue underwent permeabilization with permeabilization enzyme (24 minutes), reverse transcription and second strand synthesis performed on the slide.The following library construction was performed as per manufacturer's instructions including fragmentation, end repair & A-tailing,  post fragmentation end repair & A-tailing double sided size selection(SPRIselect), adaptor ligation, post ligation cleanup, sample Index PCR, SPRIselect. The libraries were tested using the Agilent TapeStation. The pooled libraries of each sample (100 million reads per tissue) ran on one lane of NovaSeq S4 following manufacturer's recommendations.