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Related Experiment Video

Updated: Feb 3, 2026

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A siRNA Mediated Screen During C2C12 Myogenesis.

Alexis Parenté1, Luce Pèrié1, Laetitia Magnol1

  • 1INRA, PEIRENE EA7500, USC1061 GAMAA, Université de Limoges, Limoges, France.

Methods in Molecular Biology (Clifton, N.J.)
|October 28, 2018
PubMed
Summary

This study developed a high-throughput RNA interference screen to identify genes regulating muscle growth. The method efficiently analyzes cellular phenotypes to uncover molecular mechanisms controlling myogenesis.

Keywords:
C2C12DifferentiationFunctional screenImmunostainingMyogenesisProliferationQuantitative-imaging analysissiRNA

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Area of Science:

  • Molecular Biology
  • Cell Biology
  • Genetics

Background:

  • Myogenesis is crucial for muscle development, growth, and regeneration, involving complex molecular regulation of myoblast proliferation and differentiation.
  • Understanding the intricate molecular mechanisms governing myogenesis is essential for advancing muscle biology and regenerative medicine.

Purpose of the Study:

  • To provide new insights into the molecular mechanisms and interactions regulating myogenesis.
  • To develop and implement an efficient RNA interference (RNAi) methodology for high-throughput screening.
  • To identify genes involved in signaling pathways that control muscle growth.

Main Methods:

  • A high-throughput siRNA screen was conducted using C2C12 murine myoblast cells.
  • The protocol involved analyzing a large number of cells/myotubes to detect subtle phenotypes during proliferation and fusion.
  • Quantitative assessment of cellular density, myotube quantity, fusion index, and myotube size/morphology using DAPI and myosin fluorescent markers.

Main Results:

  • The developed protocol enables the detection and quantification of both clear and subtle cellular phenotypes.
  • The use of fluorescent markers (DAPI and myosin) facilitated detailed analysis of nuclei and myotubes.
  • Four key phenotypic criteria were quantitatively assessed, providing a comprehensive evaluation of myogenesis.

Conclusions:

  • The high-throughput siRNA screen is an efficient methodology for dissecting the molecular regulation of myogenesis.
  • This approach offers a robust platform for identifying novel genes and pathways controlling muscle growth and regeneration.
  • The protocol enhances the understanding of molecular factors influencing myoblast behavior and muscle formation.