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CRISPR/Cas9 对小鼠和人造血祖细胞的高效基因破坏

JoVE Journal
Genética

This content is Free Access.

JoVE Journal Genética

Highly Efficient Gene Disruption of Murine and Human Hematopoietic Progenitor Cells by CRISPR/Cas9

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CRISPR/Cas9 对小鼠和人造血祖细胞的高效基因破坏

DOI:

10.3791/57278-v

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08:27 min

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April 10, 2018

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Lorenzo Brunetti*^1,2,3, Michael C. Gundry*^1,2,4, Ayumi Kitano, Daisuke Nakada^2,4, Margaret A. Goodell^2,4,5

¹Stem Cells & Regenerative Medicine Center,Baylor College of Medicine, ²Center for Cell and Gene Therapy,Baylor College of Medicine, ³Centro di Ricerca Emato-Oncologica (CREO),University of Perugia, ⁴Department of Molecular & Human Genetics,Baylor College of Medicine, ⁵Texas Children’s Hospital & Houston Methodist Hospital

Capítulos

00:05Título
01:05sgRNA Fwd Primer Design, DNA Template Synthesis, and ln Vitro Transcription of sgRNA
03:38Cas9-sgRNA Complexing and Electroporation
05:45Results: Use of Cas9-sgRNA Electroporation Protocol to Study Efficient Knockout of Target Gene
07:48Conclusion

Summary

Tadução automática

本文介绍了一种快速 CRISPR/Cas9-mediated 基因在小鼠和人原发性造血细胞中的破坏的协议。Cas9-sgRNA ribonucleoproteins 是通过电穿孔与 sgRNAs 产生通过体外转录和商业 Cas9。在有限的时间和财务成本下实现了高编辑效率。

Tags

CRISPR/Cas9 Gene Disruption Hematopoietic Progenitor Cells Ribonucleoprotein Single-guide RNA PCR T7 RNA Polymerase Gene Knockout Primary Cells High Efficiency Fast Turnaround Cost-effective Normal And Malignant Hematopoiesis Primary T-cells Hematopoietic Cell Lines Primary Leukemia Samples

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