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Using a Fluorescent PCR-capillary Gel Electrophoresis Technique to Genotype CRISPR/Cas9-mediated Knockout Mutants in a High-throughput Format
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遗传学
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JoVE 杂志 遗传学
Using a Fluorescent PCR-capillary Gel Electrophoresis Technique to Genotype CRISPR/Cas9-mediated Knockout Mutants in a High-throughput Format

Using a Fluorescent PCR-capillary Gel Electrophoresis Technique to Genotype CRISPR/Cas9-mediated Knockout Mutants in a High-throughput Format

DOI:
10.3791/55586-v

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08:25 min

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April 08, 2017

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, , ,
1Cancer & Stem Cell Biology Programme,Duke-NUS Medical School

Chapters

  • 00:05Title
  • 00:55Sample Preparation and Amplification of CRISPER/Cas9 Regions with Fluorescent PCR
  • 04:32Capillary Gel Electrophoresis and Electropherogram Analysis
  • 06:25Results: Genotyping HEPG2 Targeted with CRISPER/Cas9 Construct Against NAP1L1
  • 07:27Conclusion

Summary

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The genotyping technique described here, which couples fluorescent polymerase chain reaction (PCR) to capillary gel electrophoresis, allows for high-throughput genotyping of nuclease-mediated knockout clones. It circumvents limitations faced by other genotyping techniques and is more cost effective than sequencing methods.

Tags

CRISPR/Cas9Knockout MutantsGenotypingFluorescent PCRCapillary Gel ElectrophoresisHigh-throughputCell CultureCloningCell LysisPCR

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