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JoVE Journal

Developmental Biology

This content is Free Access.

CRISPR/Cas9-Mediated Highly Efficient Gene Targeting in Embryonic Stem Cells for Developing Gene-Manipulated Mouse Models

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10:57 min

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August 24th, 2022

August 24th, 2022

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3,434 Views

0:04

Introduction

1:06

Gene Targeting of Embryonic Stem Cells (ESCs)

4:16

PCR Genotyping of Targeted ESCs

7:02

Preparation of Eight-Cell Stage Embryo and Microinjection of ESCs

9:49

Results: Screening of Targeted ESC Clones

10:10

Conclusion

Transcrição

Here we present a protocol for developing genetically modified mouse models using embryonic stem cells, especially for large DNA knock-in (KI). This protocol is tuned up using CRISPR/Cas9 genome editing, resulting in significantly improved KI efficiency compared with the conventional homologous recombination-mediated linearized DNA targeting method.

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