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Preparation and Cryo-FIB micromachining of Saccharomyces cerevisiae for Cryo-Electron Tomography
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Biochemistry
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JoVE Journal Biochemistry
Preparation and Cryo-FIB micromachining of Saccharomyces cerevisiae for Cryo-Electron Tomography

Preparation and Cryo-FIB micromachining of Saccharomyces cerevisiae for Cryo-Electron Tomography

DOI:
10.3791/62351-v

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09:06 min

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November 20, 2021

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, , ,
1Central European Institute of Technology,Masaryk University

Summary

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We present a protocol for lamella preparation of plunge frozen biological specimens by cryo-focused ion beam micromachining for high-resolution structural studies of macromolecules in situ with cryo-electron tomography. The presented protocol provides guidelines for the preparation of high-quality lamellae with high reproducibility for structural characterization of macromolecules inside the Saccharomyces cerevisiae.

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Keywords: Saccharomyces CerevisiaeCryo-electron TomographyCell CultivationSterile ConditionsLaminar Flow BoxYeast Extract MediumGlucoseExponential PhaseOD MeasurementGlycerolPlasma CleaningGlow DischargingVitrobotBlottingLiquid EthaneVitrification

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