EMD-5490

Helical reconstruction
12.8 Å
EMD-5490 Deposition: 01/09/2012
Map released: 03/10/2012
Last modified: 26/03/2014
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Overview 3D View Sample Experiment Validation Volume Browser Additional data Links

EMD-5490

Reconstruction of the Wild-type Ndc80 Bonsai Decorated Microtubule on CCD for Difference Map Calculation

EMD-5490

Helical reconstruction
12.8 Å
EMD-5490 Deposition: 01/09/2012
Map released: 03/10/2012
Last modified: 26/03/2014
Overview 3D View Sample Experiment Validation Volume Browser Additional data Links
Method: Helical reconstruction
Aggregation State: Filament
Specimen preparation [1]
Concentration: 0.25 mg/mL
Buffer
pH: 6.8
Details: 80mM PIPES, 1mM MgCl2, 1mM EGTA, 1mM DTT, 0.05% Nonidet P-40, 20uM taxol
Grid
Details: C-flat 1.2/1.3
Vitrification
Cryogen name: ETHANE
Chamber humidity: 100%
Instrument: FEI VITROBOT MARK II
Method: 2 uL of 0.25 mg/mL MTs applied to grid for 1 minute, 4 uL of 0.7 mg/mL Ndc80 bonsai added, manually blot 1 minute, another 4 uL of Ndc80 applied for 1 minute, 2 uL removed with pipettor, blot for 2 seconds before plunging, 0 mm offset
Microscopy [1]
Microscope: FEI TECNAI F20
Illumination mode: FLOOD BEAM
Imaging mode: BRIGHT FIELD
Electron source: FIELD EMISSION GUN
Acceleration voltage: 120 kV
Nominal CS: 2.2 mm
Nominal defocus: 0.8 µm - 3.2 µm
Nominal magnification: 80000.0
Specimen holder model: GATAN LIQUID NITROGEN
Specimen holder details: Side-entry
Alignment procedure: LEGACY (Astigmatism: objective lens astigmatism corrected at 100Kx mag, Electron beam tilt params: )
Image Recording [1]
Detector category: CCD
Detector model: GATAN ULTRASCAN 4000 (4k x 4k)
Sampling interval: 15 µm
Number of real images: 202
Average electron dose per image: 20 e/Å2
Image processing [1]
Details: The phase-flipped particles were aligned using IHRSR in EMAN2/SPARX.
Final reconstruction
Resolution: 12.8 Å ( BY AUTHOR)
Resolution method: FSC 0.5 CUT-OFF
Algorithm: OTHER
Details: Particles were aligned using multi-model IHRSR protocol in EMAN2/SPARX with naked 13 and 14 protofilament microtubules as references. The deposited map is a segmented region for difference map calculation. No amplitude scaling was applied.
Applied Symmetry
ΔΖ: 9.04016 Å
ΔΦ: 25.76246°
Software [1]
Name Version Details
EMAN2/SPARX - -
CTF correction
Details:Phase-flipping each image
Map
Format: CCP4
Data type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation details: Reconstruction of a 14 protofilament microtubule with wild-type ndc80 Bonsai
Details: ::::EMDATABANK.org::::EMD-5490::::
Geometry
X Y Z
Dimensions 70 70 70
Origin 0 0 0
Spacing 70 70 70
Voxel size 2.74 Å 2.74 Å 2.74 Å
Contour list
Primary Level Source
True 1.67 AUTHOR