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Direct Gene Knock-out of Axolotl Spinal Cord Neural Stem Cells via Electroporation of CAS9 Protein-gRNA Complexes
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Genetics
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JoVE Journal Genetics
Direct Gene Knock-out of Axolotl Spinal Cord Neural Stem Cells via Electroporation of CAS9 Protein-gRNA Complexes

Direct Gene Knock-out of Axolotl Spinal Cord Neural Stem Cells via Electroporation of CAS9 Protein-gRNA Complexes

DOI:
10.3791/59850-v

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09:10 min

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July 09, 2019

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, , , ,
1School of Life Sciences,South China Normal University, 2The Research Institute of Molecular Pathology (IMP),Vienna Biocenter (VBC), 3Institute for Brain Research and Rehabilitation (IBRR),South China Normal University

Chapters

  • 00:04Title
  • 01:16CAS9-gRNA Ribonucleoprotein Mixture and Agarose Plate Preparation
  • 02:33Electroporator Configuration and Glass Microinjection Capillary Loading
  • 04:09Pneumatic Pump Configuration
  • 04:37CAS9-gRNA Ribonucleoprotein Mixture Injection
  • 06:05Electroporation
  • 06:45Knock-Out Efficiency Assessment
  • 07:25Results: Representative Knock-Out Effects in Axolotl Spinal Cord Neural Stem Cells
  • 08:33Conclusion

Summary

Automatic Translation

Automatic Translation

Presented here is a protocol to perform time- and space-restricted gene knock-out in axolotl spinal cords by injecting CAS9-gRNA complex into the spinal cord central canal followed by electroporation.

Tags

AxolotlSpinal CordNeural Stem CellsGene Knock-outElectroporationCAS9GRNARegenerationMolecular MechanismsNeural Stem Cell FunctionSpinal Cord InjuryAgarose Electroporation PlatePoring PulseTransfer Pulse

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