ELECTRON MICROSCOPY

Sample

Human ORC subunits (1-5) and Human CDC6 protein complex with 60bp DNA

Specimen Preperation
Sample Aggregation State PARTICLE
Vitrification Instrument LEICA EM GP
Cryogen Name ETHANE
Sample Vitrification Details The sample was applied to a glow-discharged Quantifoil R 0.6/1 300 mesh copper grid (previously washed with ethyl acetate), incubated for 10 seconds, blotted for 3 seconds, and plunged into liquid ethane using a Leica Automatic Plunge Freezer EM GP2.
3D Reconstruction
Reconstruction Method SINGLE PARTICLE
Number of Particles 130819
Reported Resolution (Å) 2.8
Resolution Method FSC 0.5 CUT-OFF
Other Details ?
Refinement Type
Symmetry Type POINT
Map-Model Fitting and Refinement
ID 1
Refinement Space REAL
Refinement Protocol FLEXIBLE FIT
Refinement Target ?
Overall B Value ?
Fitting Procedure ?
Details Initial local fitting was done using Chimera, and then Coot was used for flexible fitting and model building.
Data Acquisition
Detector Type GATAN K3 BIOQUANTUM (6k x 4k)
Electron Dose (electrons/Å2) 43.2
Imaging Experiment
Date of Experiment ?
Temprature (Kelvin)
Microscope Model TFS KRIOS
Minimum Defocus (nm) 600
Maximum Defocus (nm) 2200
Minimum Tilt Angle (degrees) ?
Maximum Tilt Angle (degrees) ?
Nominal CS ?
Imaging Mode BRIGHT FIELD
Specimen Holder Model FEI TITAN KRIOS AUTOGRID HOLDER
Nominal Magnification 105000
Calibrated Magnification ?
Source FIELD EMISSION GUN
Acceleration Voltage (kV) 300
Imaging Details ?
Imaging Experiment
Task Software Package Version
PARTICLE SELECTION Warp 1.0.9
PARTICLE SELECTION cryoSPARC 4.7.1
IMAGE ACQUISITION Warp 1.0.9
IMAGE ACQUISITION EPU ?
CTF CORRECTION Warp 1.0.9
MODEL FITTING Coot 0.9.8.95
INITIAL EULER ASSIGNMENT cryoSPARC 4.7.1
FINAL EULER ASSIGNMENT cryoSPARC 4.7.1
CLASSIFICATION cryoSPARC 4.7.1
RECONSTRUCTION cryoSPARC 4.7.1
MODEL REFINEMENT PHENIX 1.21.2_5419
Image Processing
CTF Correction Type CTF Correction Details Number of Particles Selected Particle Selection Details
PHASE FLIPPING AND AMPLITUDE CORRECTION CTF correction was done first in WARP during exposure/micrograph pre-processing, and re-corrected during the final refinements/reconstruction of the map in cryoSPARC
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