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- EMDB-5485: Bovine rotavirus DLP -

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Basic information

Entry
Database: EMDB / ID: 5485
TitleBovine rotavirus DLP
Map dataIcosahedral reconstruction of rotavirus double-layered particle using the DE-12 direct electron detector without movie frame alignment
SampleRotavirus double-layered particle:
virus
Keywordsvirus / assembly intermediate
SourceBovine rotavirus
Methodsingle particle reconstruction / cryo EM / 7.5 Å resolution
AuthorsCampbell MG / Cheng A / Brilot AF / Moeller A / Lyumkis D / Veesler D / Pan J / Harrison SC / Potter CS / Carragher B / Grigorieff N
CitationJournal: Structure / Year: 2012
Title: Movies of ice-embedded particles enhance resolution in electron cryo-microscopy.
Authors: Melody G Campbell / Anchi Cheng / Axel F Brilot / Arne Moeller / Dmitry Lyumkis / David Veesler / Junhua Pan / Stephen C Harrison / Clinton S Potter / Bridget Carragher / Nikolaus Grigorieff
Abstract: Low-dose images obtained by electron cryo-microscopy (cryo-EM) are often affected by blurring caused by sample motion during electron beam exposure, degrading signal especially at high resolution. We ...Low-dose images obtained by electron cryo-microscopy (cryo-EM) are often affected by blurring caused by sample motion during electron beam exposure, degrading signal especially at high resolution. We show here that we can align frames of movies, recorded with a direct electron detector during beam exposure of rotavirus double-layered particles, thereby greatly reducing image blurring caused by beam-induced motion and sample stage instabilities. This procedure increases the efficiency of cryo-EM imaging and enhances the resolution obtained in three-dimensional reconstructions of the particle. Using movies in this way is generally applicable to all cryo-EM samples and should also improve the performance of midrange electron microscopes that may have limited mechanical stability and beam coherence.
DateDeposition: Aug 24, 2012 / Header (metadata) release: Sep 12, 2012 / Map release: Sep 12, 2012 / Last update: Nov 21, 2012

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.1
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.1
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

Fileemd_5485.map.gz (map file in CCP4 format, 2000001 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
800 pix
1.42 Å/pix.
= 1139.2 Å
800 pix
1.42 Å/pix.
= 1139.2 Å
800 pix
1.42 Å/pix.
= 1139.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.424 Å
Density
Contour Level:0.1 (by author), 0.1 (movie #1):
Minimum - Maximum-0.15407689 - 0.24396071
Average (Standard dev.)-0.02877968 (0.0324422)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions800800800
Origin000
Limit799799799
Spacing800800800
CellA=B=C: 1139.2001 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.4241.4241.424
M x/y/z800800800
origin x/y/z0.0000.0000.000
length x/y/z1139.2001139.2001139.200
α/β/γ90.00090.00090.000
start NX/NY/NZ-150-1500
NX/NY/NZ301301151
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS800800800
D min/max/mean-0.1540.244-0.029

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Supplemental data

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Sample components

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Entire Rotavirus double-layered particle

EntireName: Rotavirus double-layered particle / Details: The sample was monodisperse. / Number of components: 1
Oligomeric State: 12 x VP1, 120 x VP2, 12 x VP3, 780 x VP6, 11 x RNA
MassTheoretical: 70 MDa

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Component #1: virus, Bovine rotavirus

VirusName: Bovine rotavirus / Class: VIRUS-LIKE PARTICLE / Empty: No / Enveloped: No / Isolate: STRAIN
MassTheoretical: 70 MDa
SpeciesSpecies: Bovine rotavirus
Source (natural)Host Species: Bos primigenius / Host category: VERTEBRATES
Shell #1Name of element: DLP / Diameter: 700 Å / T number(triangulation number): 13

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionSpecimen conc.: 5 mg/ml / pH: 7.4
Support film1.2-1.3 C-flat
VitrificationInstrument: FEI VITROBOT MARK II / Cryogen name: ETHANE / Temperature: 120 K / Humidity: 85 % / Method: Blot for 7 seconds before plunging

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Electron microscopy imaging

Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company
ImagingMicroscope: FEI TECNAI F20 / Date: Nov 28, 2011
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Electron dose: 32 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 42135 X (calibrated) / Cs: 2 mm / Imaging mode: BRIGHT FIELD / Defocus: 1000 - 2400 nm
Specimen HolderModel: GATAN LIQUID NITROGEN
CameraDetector: DIRECT ELECTRON DE-12 (4k x 3k)

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Image acquisition

Image acquisition #1
501
Image acquisition #1~
6
Image acquisition #1~~
Moviesreco rdedat25fr amessecond

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Image processing

ProcessingMethod: single particle reconstruction
Details: Manual particle selection, refinement using Frealign.
Number of projections: 807 / Applied symmetry: I (icosahedral)
3D reconstructionAlgorithm: Projection matching / Software: Frealign / CTF correction: Each particle
Details: Final map was calculated from 807 particles with icosahedral symmetry imposed
Resolution: 7.5 Å / Resolution method: FSC 0.143

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