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Nat. Commun., Sep 16 2024, 15 (1) | https://doi.org/10.1038/s41467-024-52493-1
Highly efficient CRISPR-mediated genome editing through microfluidic droplet cell mechanoporation
You-Jeong Kim1,2,7, Dayoung Yun3,7, Jungjoon K. Lee4, Cheulhee Jung3 & Aram J. Chung1,2,5,6
1Department of Bioengineering, Korea University, Seoul, Republic of Korea.
2Interdisciplinary Program in Precision Public Health (PPH), Korea University, Seoul, Republic of Korea.
3Department of Biotechnology, Korea University, Seoul, Republic of Korea.
4Department of Biochemistry, National University of Singapore, Singapore, Singapore.
5School of Biomedical Engineering, Korea University, Seoul, Republic of Korea.
6MxT Biotech, Seoul, Republic of Korea.
7These authors contributed equally: You-Jeong Kim, Dayoung Yun.
Corresponding authors
Correspondence to Cheulhee Jung or Aram J. Chung.
Abstract
Clustered regularly interspaced short palindromic repeats (CRISPR)-based editing tools have transformed the landscape of genome editing. However, the absence of a robust and safe CRISPR delivery method continues to limit its potential for therapeutic applications. Despite the emergence of various methodologies aimed at addressing this challenge, issues regarding efficiency and editing operations persist. We introduce a microfluidic gene delivery system, called droplet cell pincher (DCP), designed for highly efficient and safe genome editing. This approach combines droplet microfluidics with cell mechanoporation, enabling encapsulation and controlled passage of cells and CRISPR systems through a microscale constriction. Discontinuities created in cell and nuclear membranes upon passage facilitate the rapid CRISPR-system internalization into the nucleus. We demonstrate the successful delivery of various macromolecules, including mRNAs (~98%) and plasmid DNAs (~91%), using this platform, underscoring the versatility of the DCP and leveraging it to achieve successful genome engineering through CRISPR-Cas9 delivery. Our platform outperforms electroporation, the current state-of-the-art method, in three key areas: single knockouts (~6.5-fold), double knockouts (~3.8-fold), and knock-ins (~3.8-fold). These results highlight the potential of our platform as a next-generation tool for CRISPR engineering, with implications for clinical and biological cell-based research.
논문정보
- Research article
- 2024년 09월 (BRIC 등록일 2024-09-19)
- 국내(교신)+국외 연구진
- 바이오·의료융합 > 바이오·의료융합 기타
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