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Related Experiment Video

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Transcriptome Analysis of Single Cells
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Live Cell Genomics: Cell-Specific Transcriptome Capture in Live Single Cells from Complex Tissues.

Jaehee Lee1, James Eberwine2

  • 1Department of Systems Pharmacology and Translational Therapeutics, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA.

Methods in Molecular Biology (Clifton, N.J.)
|November 12, 2021
PubMed
Summary

We developed transcriptome in vivo analysis (TIVA) to isolate single-cell mRNA from live tissues. This noninvasive method uses TIVA tags and cell-penetrating peptides for accurate transcriptome profiling in the natural cellular environment.

Keywords:
Cell-penetrating peptidePhotoactivationSingle-cell transcriptomeTranscriptome in vivo analysisTranscriptomics

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Area of Science:

  • Molecular Biology
  • Genomics
  • Cell Biology

Background:

  • Single-cell transcriptome analysis is crucial for understanding cellular heterogeneity in health and disease.
  • Accurate isolation of poly-A RNA from individual cells is a prerequisite for transcriptome profiling.
  • Existing methods face challenges in preserving cellular context and achieving noninvasive isolation.

Purpose of the Study:

  • To develop a novel method for accurate, noninvasive isolation of single-cell transcriptomes from live tissues.
  • To enable the characterization of cell-specific transcriptomes within their native microenvironment.

Main Methods:

  • Development of transcriptome in vivo analysis (TIVA), a procedure utilizing a TIVA tag for RNA capture.
  • Delivery of the TIVA tag into the cytoplasm of live cells via cell-penetrating peptides (CPPs).
  • Photoactivation of the TIVA tag to bind mRNA within the cellular cytoplasm.

Main Results:

  • TIVA enables the capture of the transcriptome from selected cells in their natural microenvironment.
  • The combination of CPPs and photoactivation provides a noninvasive method for mRNA isolation.
  • This approach allows for accurate isolation of single-cell mRNA from live cells within tissues.

Conclusions:

  • Transcriptome in vivo analysis (TIVA) is a pioneering noninvasive technique for single-cell mRNA isolation.
  • TIVA facilitates the accurate characterization of cell-specific transcriptomes in their native tissue context.
  • This method advances the field of single-cell analysis, offering new insights into cellular function and disease.