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Stable Knockdown of Genes Encoding Extracellular Matrix Proteins in the C2C12 Myoblast Cell Line Using Small-Hairpin (sh)RNA
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JoVE Journal Genetics
Stable Knockdown of Genes Encoding Extracellular Matrix Proteins in the C2C12 Myoblast Cell Line Using Small-Hairpin (sh)RNA

Stable Knockdown of Genes Encoding Extracellular Matrix Proteins in the C2C12 Myoblast Cell Line Using Small-Hairpin (sh)RNA

DOI:
10.3791/60824-v

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12:19 min

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February 12, 2020

•

, ,
1Orthopaedic Research Laboratories, Leni & Peter W. May Department of Orthopaedics,Icahn School of Medicine at Mount Sinai

Chapters

  • 00:04Introduction
  • 00:55C2C12 Transfection
  • 02:35Transfected C2C12 Selection
  • 03:15C2C12 Differentiation
  • 03:59Phenotypic Analysis
  • 06:10Western Blot Analysis Sample Preparation
  • 09:37Results: Representative C2C12 Selection, Differentiation, and Knockdown Validation
  • 11:19Conclusion

Summary

Automatic Translation

Automatic Translation

We provide a protocol to stably knock down genes encoding extracellular matrix (ECM) proteins in C2C12 myoblasts using small-hairpin (sh) RNA. Targeting ADAMTSL2 as an example, we describe the methods for the validation of the knockdown efficiency on the mRNA, protein, and cellular level during C2C12 myoblast to myotube differentiation.

Tags

Keywords: C2C12 MyoblastShRNAGene KnockdownExtracellular Matrix ProteinsStable Cell LineTransfectionPuromycin SelectionCell CultureMyotube Formation

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