Using the Volta phase plate with defocus for cryo-EM single particle analysis
- Max Planck Institute of Biochemistry, Germany
- Max Planck Institute for Biophysical Chemistry, Germany
Abstract
Previously, we reported an in-focus data acquisition method for cryo-EM single particle analysis with the Volta phase plate (VPP) (Danev and Baumeister, 2016). Here, we extend the technique to include a small amount of defocus which enables contrast transfer function measurement and correction. This hybrid approach simplifies the experiment and increases the data acquisition speed. It also removes the resolution limit inherent to the in-focus method thus allowing 3D reconstructions with resolutions better than 3 Å.
Data availability
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Volta phase plate with defocus cryo-EM dataset of Thermoplasma acidophilum 20S proteasomePublicly available at the Electron Microscopy Pilot Image Archive (accession no: EMPIAR-10078).
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Thermoplasma acidophilum 20S proteasome map reconstructed with Relion 2.0 particle polishing from defocused Volta phase plate cryo-EM dataPublicly available at the EMDB Protein Data Bank in Europe (accession no: EMD-3455).
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Thermoplasma acidophilum 20S proteasome map reconstructed with MotionCor2 and Relion 2.0 from defocused Volta phase plate cryo-EM dataPublicly available at the EMDB Protein Data Bank in Europe (accession no: EMD-3456).
Article and author information
Author details
Funding
Max-Planck-Gesellschaft
- Radostin Danev
- Dimitry Tegunov
- Wolfgang Baumeister
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2017, Danev et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Using the Volta phase plate with defocus for cryo-EM single particle analysis
eLife 6:e23006.
https://doi.org/10.7554/eLife.23006
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