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哺乳动物细胞中基于CRISPR的聚集基因屏幕

JoVE Journal
Genetics

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JoVE Journal Genetics

Pooled CRISPR-Based Genetic Screens in Mammalian Cells

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哺乳动物细胞中基于CRISPR的聚集基因屏幕

DOI:

10.3791/59780-v

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00:09 min

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September 04, 2019

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Katherine Chan*¹, Amy Hin Yan Tong*¹, Kevin R Brown, Patricia Mero, Jason Moffat^2,3

¹Donnelly Centre,University of Toronto, ²Department of Molecular Genetics,University of Toronto, ³Institute for Biomaterials and Biomedical Engineering,University of Toronto

Chapters

00:04Title
01:36Pooled CRISPR sgRNA Lentiviral Library Plasmid Amplification
02:24Large-scale CRISPR sgRNA Library Lentivirus Production
03:47Primary Screen Infection, Selection, and Cell Passaging
05:41CRISPR Sample Preparation and Sequencing
06:43Results: Genome-scale CRISPR Screening
07:54Conclusion

Summary

Automatic Translation

CRISPR-Cas9技术提供了一种有效的方法，可以精确编辑任何细胞类型的哺乳动物基因组，并代表一种执行全基因组基因筛选的新方法。此处提供了一个详细的协议，讨论成功执行池全基因组 CRISPR-Cas9 屏幕所需的步骤。

Tags

Keywords: CRISPR Genetic Screens Mammalian Cells Drop-out Screens Guide Libraries Essential Genes Cancer Drug Mechanisms Cell Lines Lentivirus Antibiotic Selection Plasmid Transformation Electrocompetent Cells LB Carbenicillin Plasmid Purification 293T Cells Transfection

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