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September 12, 2025Open Access

Optimized CRISPR/Cas9 Electroporation and Single Cell Cloning Protocol for Generating Pure Cellular Models in Human Immortalized Myoblasts

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Authors

ARAdrien RihouxAGAlyssa GagneJMJean Mezreani

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Overview

This work demonstrates optimized electroporation and cloning for CRISPR/Cas9 in human myoblasts, highlighting efficacy and protocol applicability.

Key Points

  • An 84% success rate for IARS1 knockout was achieved using the optimized CRISPR/Cas9 protocol for human myoblasts.
  • Low confluency during electroporation increased both editing rates and clonal outgrowth, enhancing model generation.
  • High-resolution melting detected 96% to 100% of actual edits, streamlining Sanger sequencing requirements significantly.
  • The developed protocol is practical for generating pure edited myoblast lines, aiding in neuromuscular disease research.

Cite This Study

Rihoux et al. (2025) studied this question.

synapsesocial.com/papers/68d44f7b31b076d99fa56bc8https://doi.org/10.1101/2025.09.08.674725
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Also Consider

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  1. 1Streamlined, single-step non-viral CRISPR-Cas9 knockout strategy enhances gene editing efficiency in primary human chondrocyte populations2024 · 2 citations
  2. 2Rapid and Robust Generation of Homozygous Fluorescent Reporter Knock-In Cell Pools by CRISPR-Cas92025 · 1 citations
  3. 3Versatile electroporation protocols enable reproducible CRISPR-RNP delivery across multiple primary mouse cells of the hematopoietic lineage2026
  4. 4Nanopuncturing‐Enabled Efficient Gene Editing for Isogenic Human iPSC‐Derived Organoid Engineering2025
  5. 5Nanopuncturing‐Enabled Efficient Gene Editing for Isogenic Human iPSC‐Derived Organoid Engineering2025