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Cell-Type Specific Protein Purification and Identification from Complex Tissues Using a Mutant Methionine tRNA Synthetase Mouse Line
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Neurociência
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JoVE Journal Neurociência
Cell-Type Specific Protein Purification and Identification from Complex Tissues Using a Mutant Methionine tRNA Synthetase Mouse Line

Cell-Type Specific Protein Purification and Identification from Complex Tissues Using a Mutant Methionine tRNA Synthetase Mouse Line

DOI:
10.3791/63713-v

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07:39 min

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April 13, 2022

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, , , , , ,
1Max-Planck-Institute for Brain Research,Synaptic Plasticity Department, 2Department of Biochemistry and Molecular Biology,Veterinary School, Complutense University of Madrid

Capítulos

  • 00:05Introduction
  • 00:40Tissue Harvesting, Lysis, and Protein Extraction
  • 03:03Protein Purification
  • 05:28Results: Cell-Type-Specific Protein Labeling with Azidonorleucine
  • 06:35Conclusion

Summary

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This protocol describes how to perform cell-type-specific protein labeling with azidonorleucine (ANL) using a mouse line expressing a mutant L274G-Methionine tRNA synthetase (MetRS*) and the necessary steps for labeled cell-type-specific proteins isolation. We outline two possible ANL administration routes in live mice by (1) drinking water and (2) intraperitoneal injections.

Tags

Keywords: Cell-type SpecificProtein PurificationProtein IdentificationMutant Methionine TRNA SynthetaseCell Type Specific ProteinsIn VitroIn VivoImmunofluorescentAlkylationIodoacetamideBuffer ExchangeClick ChemistryAzidonorleucine IncorporationNeutravidin Binding

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