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Metabolic Labeling of Yeast Proteins

Bio-protocol Editor Bio-protocol Editor

Published: Jan 5, 2011 DOI: 10.21769/BioProtoc.13 Views: 13464

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Abstract

Proteins of Saccharomyces cerevisiae can be metabolically labeled with (35) methionine. After labeling, a protocol is described for the mechanical disruption of yeast cells or conversion to spheroplasts, with subsequent lysis before immunoprecipitation of the proteins.

Materials and Reagents

  1. TRAN35S-label (MP Biomedicals)
  2. Methionine
  3. FLAG peptides
  4. Immunoprecipitation buffer
  5. Digitonin (EMD Chemicals)
  6. 1x SDS
  7. Medium minus methionine (SD-Met)

Equipment

  1. Bench-top centrifuges 
  2. Charcoal filter paper
  3. 50 ml conical tube
  4. 125 ml Erlenmeyer flask with vented cap (Nalgene)
  5. O-ring cap tube
  6. Geiger counter
  7. Labeled radioactive beaker
  8. Kimwipes
  9. 1.5 ml Eppendorf tubes

Procedure

Copyright: © 2011 The Authors; exclusive licensee Bio-protocol LLC.

Category

Biochemistry > Protein > Labeling

Microbiology > Microbial biochemistry > Protein

Cell Biology > Cell viability > Cell lysis

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