Computational protocol: The structure of the COPII transport vesicle coat assembled on membranes

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Protocol publication

[…] Tomograms were aligned based on the positions of gold fiducials and reconstructed by weighted back-projection, using the IMOD () and raptor () software packages. Visual analysis and segmentation of reconstructed tomograms were performed with Amira (Visualisation Sciences Group, an FEI company) and Chimera (). Sub-tomogram averaging was carried out essentially as described previously () using Matlab scripts adapted from the TOM/AV3 tomography toolbox () and using the Dynamo package ().Two datasets collected on the FEI Titan Krios were used for sub-tomogram averaging. A ‘far from focus’ dataset collected at nominal defoci of 4–6 µm contained 16 tubes (typically 1 tube in each tomogram). A second, close to focus dataset (nominal defoci 1.5–2.5 µm), contained 26 tubes. Both unbinned datasets had a pixel size of 4.3 Å. […]

Pipeline specifications

Software tools IMOD, RAPTOR, Dynamo
Application cryo-electron tomography