Efficient spatial gene expression profiling using split-probe ligation and rolling circle amplification

Our study presents a new approach for spatial gene expression profiling based on split-probe ligation and rolling circle amplification (RCA). By hybridizing split-probes to adjacent RNA regions, enzymatically ligating them to form circular DNA templates, and amplifying these via RCA, we achieve robust in situ RNA detection and genotyping in tissue sections and whole-mount samples. Coupled with in situ sequencing, our method enabled the mapping of 82 genes in the kidneys of healthy and diabetic male and female mice, revealing distinct localization of Aqp4 in proximal tubules and principal cells of the collecting ducts, as well as sex-specific transcriptomic alterations in diabetic kidneys with high spatial resolution.