- EMDB-21953: Downsampled and filtered tomogram of a region from a cryoFIB-SEM-... -
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Open data
ID or keywords:
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Basic information
Entry
Database: EMDB / ID: EMD-21953
Title
Downsampled and filtered tomogram of a region from a cryoFIB-SEM-generated lamellae of yeast cells under heat shock stress showing large protein aggregates
Map data
Binned-by-8 and filtered tomogram of a region from a cryoFIB-SEM-generated lamellae of yeast cells under heat shock stress showing large protein aggregates.
Sample
Cell: Downsamples and filtered tomogram of a region from a cryoFIB-SEM-generated lamellae of yeast cells under heat shock stress showing large protein aggregates
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)
P01NS092525
United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
5P41GM103832
United States
National Institutes of Health/National Institute on Aging (NIH/NIA)
PO1AG054407
United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)
F32NS086253
United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
S10OD021600
United States
Department of Energy (DOE, United States)
BERFWP 100463
United States
Citation
Journal: Structure / Year: 2020 Title: Multi-scale 3D Cryo-Correlative Microscopy for Vitrified Cells. Authors: Gong-Her Wu / Patrick G Mitchell / Jesus G Galaz-Montoya / Corey W Hecksel / Emily M Sontag / Vimal Gangadharan / Jeffrey Marshman / David Mankus / Margaret E Bisher / Abigail K R Lytton- ...Authors: Gong-Her Wu / Patrick G Mitchell / Jesus G Galaz-Montoya / Corey W Hecksel / Emily M Sontag / Vimal Gangadharan / Jeffrey Marshman / David Mankus / Margaret E Bisher / Abigail K R Lytton-Jean / Judith Frydman / Kirk Czymmek / Wah Chiu / Abstract: Three-dimensional (3D) visualization of vitrified cells can uncover structures of subcellular complexes without chemical fixation or staining. Here, we present a pipeline integrating three imaging ...Three-dimensional (3D) visualization of vitrified cells can uncover structures of subcellular complexes without chemical fixation or staining. Here, we present a pipeline integrating three imaging modalities to visualize the same specimen at cryogenic temperature at different scales: cryo-fluorescence confocal microscopy, volume cryo-focused ion beam scanning electron microscopy, and transmission cryo-electron tomography. Our proof-of-concept benchmark revealed the 3D distribution of organelles and subcellular structures in whole heat-shocked yeast cells, including the ultrastructure of protein inclusions that recruit fluorescently-labeled chaperone Hsp104. Since our workflow efficiently integrates imaging at three different scales and can be applied to other types of cells, it could be used for large-scale phenotypic studies of frozen-hydrated specimens in a variety of healthy and diseased conditions with and without treatments.
Download / File: emd_21953.map.gz / Format: CCP4 / Size: 174.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation
Binned-by-8 and filtered tomogram of a region from a cryoFIB-SEM-generated lamellae of yeast cells under heat shock stress showing large protein aggregates.
Voxel size
X=Y=Z: 27.664 Å
Density
Minimum - Maximum
-2.9999998 - 2.9999998
Average (Standard dev.)
-0.00015260548 (±0.9856359)
Symmetry
Space group: 1
Details
EMDB XML:
Map geometry
Axis order
X
Y
Z
Origin
0
0
-103
Dimensions
463
479
206
Spacing
479
463
206
Cell
A: 13251.054 Å / B: 12808.43 Å / C: 5698.783 Å α=β=γ: 90.0 °
CCP4 map header:
mode
Image stored as Reals
Å/pix. X/Y/Z
27.663995824635
27.663995680346
27.663995145631
M x/y/z
479
463
206
origin x/y/z
0.000
0.000
0.000
length x/y/z
13251.054
12808.430
5698.783
α/β/γ
90.000
90.000
90.000
start NX/NY/NZ
79
74
0
NX/NY/NZ
93
103
213
MAP C/R/S
1
2
3
start NC/NR/NS
0
0
-103
NC/NR/NS
479
463
206
D min/max/mean
-3.000
3.000
-0.000
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Supplemental data
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Sample components
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Entire : Downsamples and filtered tomogram of a region from a cryoFIB-SEM-...
Entire
Name: Downsamples and filtered tomogram of a region from a cryoFIB-SEM-generated lamellae of yeast cells under heat shock stress showing large protein aggregates
Components
Cell: Downsamples and filtered tomogram of a region from a cryoFIB-SEM-generated lamellae of yeast cells under heat shock stress showing large protein aggregates
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Supramolecule #1: Downsamples and filtered tomogram of a region from a cryoFIB-SEM-...
Supramolecule
Name: Downsamples and filtered tomogram of a region from a cryoFIB-SEM-generated lamellae of yeast cells under heat shock stress showing large protein aggregates type: cell / ID: 1 / Parent: 0
Focused ion beam - Instrument: OTHER / Focused ion beam - Ion: OTHER / Focused ion beam - Voltage: 30 kV / Focused ion beam - Current: 20 nA / Focused ion beam - Duration: 60 sec. / Focused ion beam - Temperature: 100 K / Focused ion beam - Initial thickness: 1000 nm / Focused ion beam - Final thickness: 520 nm Focused ion beam - Details: The value given for _emd_sectioning_focused_ion_beam.instrument is Zeiss Crossbeam 540 FIB-SEM. This is not in a list of allowed values set(['DB235', 'OTHER']) so OTHER is ...Focused ion beam - Details: The value given for _emd_sectioning_focused_ion_beam.instrument is Zeiss Crossbeam 540 FIB-SEM. This is not in a list of allowed values set(['DB235', 'OTHER']) so OTHER is written into the XML file.
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Electron microscopy
Microscope
TFS KRIOS
Image recording
Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 0.8264 e/Å2
Electron beam
Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron optics
Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
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Image processing
Final reconstruction
Algorithm: SIMULTANEOUS ITERATIVE (SIRT) / Software - Name: IMOD Details: The SIRT, downsampled, and filtered tomogram was computed for visualization only. The related EMDB entry showcasing a ribosome subtomogram average used similar tomograms but reconstructed ...Details: The SIRT, downsampled, and filtered tomogram was computed for visualization only. The related EMDB entry showcasing a ribosome subtomogram average used similar tomograms but reconstructed with weighted-back-projection, at full-size, and unfiltered. Number images used: 121
CTF correction
Software - Name: IMOD / Details: 3D CTF correction
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