EMD-3682 Experiments and Validation

Structure of a pre-catalytic spliceosome (B1 map)

Single particle reconstruction
Overview of EMD-3682
Sample name: Pre-catalytic B complex Spliceosome
Organism: Saccharomyces cerevisiae
Related EM entries by publication: EMD-3683, EMD-3684, EMD-3685, EMD-3686, EMD-3687, EMD-3688

Map parameters

Recommended contour level: 0.038 (author)
Number of grid points: 480 × 480 × 480
Voxel size: 1.43 × 1.43 × 1.43 Å
Minimum density: -0.072
Maximum density: 0.162
Average density: -0.00
Standard deviation: 0.003

Sample information

Sample name: Pre-catalytic B complex Spliceosome
Protein: Pre-catalytic B complex Spliceosome

Validation information

Experimental information

 

Image processing

Applied symmetry: C1
Software: RELION
Number of particles: 254095
Details: A temperature factor of -125 A2 was applied (sharpened B1 map). Movies were binned once and aligned using MOTIONCORR.

Imaging

Session
Microscope model: FEI TITAN KRIOS
Electron source: FIELD EMISSION GUN
Electron dose (e/A**2): 56.0
Energy filter: GIF Quantum
Nominal CS (mm): 2.70
Nominal magnification: 81000.0
Holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Acquisition
Number of images: 5115
Sampling interval (μm): 5.0

Specimen

Preparation
Specimen state: particle
Specimen concentration (mg/mL): 1.50
Buffer: Buffer pH: HEPES, 7.9; EDTA, 8.0
Vitrification
Apparatus: FEI VITROBOT MARK III
Cryogen: ETHANE
Humidity: 100.0
Details: Grids were glow-discharged for 15 s before deposition of 3 microliter sample (~1.5 mg mL-1), and subsequently incubated for 2-3.5 s before blotting and vitrification by plunging into liquid ethane with a Vitrobot Mark III (FEI) operated at 4 degrees Celsius and 100% humidity.