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Efficient Spatial Gene Expression Profiling Using Split-Probe Ligation and Rolling Circle Amplification.

Xueqian Xia1, Zhaoxiang Xie1, Yu Yang1

  • 1School of Medicine, Huaqiao University, Xiamen, Fujian, 361021, China.

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|December 3, 2025
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Summary
This summary is machine-generated.

A new in situ RNA imaging method uses split-probe ligation and rolling circle amplification (RCA) for spatial gene expression profiling. This technique successfully mapped gene patterns in mouse kidneys, revealing sex-specific changes in diabetes.

Keywords:
RNA imagingRNA in situ hybridizationin situ sequencingrolling circle amplificationspatial transcriptomics

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Area of Science:

  • Molecular Biology
  • Genomics
  • Biotechnology

Background:

  • Spatial transcriptomics enables high-resolution mapping of RNA within native cellular environments.
  • Current methods capture or image RNA in situ, preserving spatial context for gene regulation studies.

Purpose of the Study:

  • To introduce a novel in situ RNA imaging method for spatial gene expression profiling.
  • To demonstrate robust RNA detection and genotyping in various tissue types.
  • To map spatial gene expression patterns in healthy and diabetic mouse kidneys.

Main Methods:

  • Developed an in situ RNA imaging technique utilizing split-probe ligation and rolling circle amplification (RCA).
  • Split-probes hybridize to target RNA, ligate, form circular DNA templates, and amplify via RCA for signal enhancement.
  • Coupled the imaging technique with in situ sequencing to profile 82 genes.

Main Results:

  • The method achieved robust in situ RNA detection and genotyping in tissue sections and whole-mount samples.
  • Spatial expression patterns of 82 genes were mapped in mouse kidneys.
  • Distinct localization of Aqp4 was observed in proximal tubules and collecting ducts.
  • Sex-specific transcriptomic alterations in diabetic kidneys were uncovered with spatial resolution.

Conclusions:

  • The split-probe ligation and RCA-based method provides a powerful tool for spatial gene expression analysis.
  • This technique offers high-resolution insights into tissue architecture and cellular function.
  • The findings highlight sex-specific kidney changes in diabetes, paving the way for targeted research.