+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-18186 | |||||||||
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Title | C. elegans L1 larva | |||||||||
Map data | Bin4 tomogram weighted backprojection | |||||||||
Sample |
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Keywords | Caenorhabditis / elegans / touch receptor cell / microtubule / L1 / larva / C. elegans / STRUCTURAL PROTEIN | |||||||||
Biological species | Caenorhabditis elegans (invertebrata) | |||||||||
Method | electron tomography / cryo EM | |||||||||
Authors | Schioetz OH / Kaiser CJO / Klumpe S / Klebl DP / Schneider J / Beck F / Plitzko JM | |||||||||
Funding support | Germany, 1 items
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Citation | Journal: Nat Methods / Year: 2023 Title: Serial Lift-Out: sampling the molecular anatomy of whole organisms. Authors: Oda Helene Schiøtz / Christoph J O Kaiser / Sven Klumpe / Dustin R Morado / Matthias Poege / Jonathan Schneider / Florian Beck / David P Klebl / Christopher Thompson / Jürgen M Plitzko / Abstract: Cryo-focused ion beam milling of frozen-hydrated cells and subsequent cryo-electron tomography (cryo-ET) has enabled the structural elucidation of macromolecular complexes directly inside cells. ...Cryo-focused ion beam milling of frozen-hydrated cells and subsequent cryo-electron tomography (cryo-ET) has enabled the structural elucidation of macromolecular complexes directly inside cells. Application of the technique to multicellular organisms and tissues, however, is still limited by sample preparation. While high-pressure freezing enables the vitrification of thicker samples, it prolongs subsequent preparation due to increased thinning times and the need for extraction procedures. Additionally, thinning removes large portions of the specimen, restricting the imageable volume to the thickness of the final lamella, typically <300 nm. Here we introduce Serial Lift-Out, an enhanced lift-out technique that increases throughput and obtainable contextual information by preparing multiple sections from single transfers. We apply Serial Lift-Out to Caenorhabditis elegans L1 larvae, yielding a cryo-ET dataset sampling the worm's anterior-posterior axis, and resolve its ribosome structure to 7 Å and a subregion of the 11-protofilament microtubule to 13 Å, illustrating how Serial Lift-Out enables the study of multicellular molecular anatomy. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_18186.map.gz | 2.7 GB | EMDB map data format | |
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Header (meta data) | emd-18186-v30.xml emd-18186.xml | 13.4 KB 13.4 KB | Display Display | EMDB header |
Images | emd_18186.png | 317.9 KB | ||
Filedesc metadata | emd-18186.cif.gz | 4.3 KB | ||
Others | emd_18186_additional_1.map.gz | 276.9 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-18186 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-18186 | HTTPS FTP |
-Related structure data
Related structure data | C: citing same article (ref.) |
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-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_18186.map.gz / Format: CCP4 / Size: 2.9 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | Bin4 tomogram weighted backprojection | ||||||||||||||||||||
Voxel size | X=Y=Z: 7.56 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Additional map: #1
File | emd_18186_additional_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : C. elegans L1 larva, CGC strain NK2476
Entire | Name: C. elegans L1 larva, CGC strain NK2476 |
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Components |
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-Supramolecule #1: C. elegans L1 larva, CGC strain NK2476
Supramolecule | Name: C. elegans L1 larva, CGC strain NK2476 / type: tissue / ID: 1 / Parent: 0 |
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Source (natural) | Organism: Caenorhabditis elegans (invertebrata) / Strain: NK2476 / Tissue: Whole L1 larva |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | electron tomography |
Aggregation state | tissue |
-Sample preparation
Buffer | pH: 7.5 / Details: M9 buffer + 20% Ficoll 400 |
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Grid | Model: Homemade / Material: COPPER / Mesh: 50 / Support film - Material: FORMVAR / Support film - topology: CONTINUOUS / Support film - Film thickness: 100 |
Vitrification | Cryogen name: NITROGEN / Details: High pressure freezing, Leica EM ICE. |
Details | Developmentally arrested L1 larvae |
High pressure freezing | Instrument: OTHER Details: The value given for _em_high_pressure_freezing.instrument is Leica EM ICE. This is not in a list of allowed values {'LEICA EM PACT2', 'BAL-TEC HPM 010', 'OTHER', 'LEICA EM HPM100', 'LEICA EM ...Details: The value given for _em_high_pressure_freezing.instrument is Leica EM ICE. This is not in a list of allowed values {'LEICA EM PACT2', 'BAL-TEC HPM 010', 'OTHER', 'LEICA EM HPM100', 'LEICA EM PACT', 'EMS-002 RAPID IMMERSION FREEZER'} so OTHER is written into the XML file. |
Cryo protectant | 20% Ficoll 400 |
Sectioning | Focused ion beam - Instrument: OTHER / Focused ion beam - Ion: OTHER / Focused ion beam - Voltage: 30 / Focused ion beam - Current: 0.03 / Focused ion beam - Duration: 1800 / Focused ion beam - Temperature: 80 K / Focused ion beam - Initial thickness: 4000 / Focused ion beam - Final thickness: 250 Focused ion beam - Details: Serial Lift-Out of 160000 nm original volume sectioned into 4000 nm slices, thinned to roughly 250 nm. The value given for _em_focused_ion_beam.instrument is TFS Aquilos 2. ...Focused ion beam - Details: Serial Lift-Out of 160000 nm original volume sectioned into 4000 nm slices, thinned to roughly 250 nm. The value given for _em_focused_ion_beam.instrument is TFS Aquilos 2. This is not in a list of allowed values {'DB235', 'OTHER'} so OTHER is written into the XML file. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 4.5 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 64000 |
Specialist optics | Energy filter - Name: TFS Selectris X / Energy filter - Slit width: 10 eV |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Image recording | Film or detector model: FEI FALCON IV (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 41 / Average exposure time: 0.73 sec. / Average electron dose: 3.23 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Final reconstruction | Algorithm: BACK PROJECTION / Software - Name: IMOD (ver. 4.12.32) / Details: ARETOMO 1.3.3 tilt series alignment / Number images used: 41 |
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