Structures and distributions of SARS-CoV-2 spike proteins on intact virions

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1. Version published to 10.1038/s41586-020-2665-2

   Aug 17, 2020
2. 

   ScreenIT

   Jul 2, 2020

   SciScore for 10.1101/2020.06.27.174979: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   NIH rigor criteria are not applicable to paper type.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Next, the membranes were washed 3 times in PBS-T, incubated with HRP conjugated anti-mouse antibodies for 1 h, washed again 3 times in PBS-T, incubated with western Lightning Plus-ECL reagent (Perkin Elmer; Waltham, MA) and imaged using an Intas ChemoCam Imager 3.2 (Intas, Göttingen).	anti-mouse suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Cells and Virus: VeroE6 cells were obtained from ATCC and were cultured in Dulbecco’s modified Eagle medium (DMEM, Life Technologies) containing 10% fetal bovine serum, 100 U/mL penicillin, 100 μg/mL streptomycin and 1% non-essential amino acids …

   SciScore for 10.1101/2020.06.27.174979: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   NIH rigor criteria are not applicable to paper type.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Next, the membranes were washed 3 times in PBS-T, incubated with HRP conjugated anti-mouse antibodies for 1 h, washed again 3 times in PBS-T, incubated with western Lightning Plus-ECL reagent (Perkin Elmer; Waltham, MA) and imaged using an Intas ChemoCam Imager 3.2 (Intas, Göttingen).	anti-mouse suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Cells and Virus: VeroE6 cells were obtained from ATCC and were cultured in Dulbecco’s modified Eagle medium (DMEM, Life Technologies) containing 10% fetal bovine serum, 100 U/mL penicillin, 100 μg/mL streptomycin and 1% non-essential amino acids (complete medium).	VeroE6 suggested: JCRB Cat# JCRB1819, RRID:CVCL_YQ49)
   Infectious supernatants containing SARS-CoV-2 virions were obtained from Calu-3 cells infected with P3 virus at an MOI = 5 for 48 h and processed as described above.	Calu-3 suggested: None
   Software and Algorithms
   Sentences	Resources
   Tomograms were low-pass filtered to 50 Å for better visualization in EMAN2.2 (36) and tomographic slices were visualized with IMOD.	IMOD suggested: (IMOD, RRID:SCR_003297)
   Extraction of S trimers from tomograms: The initial steps of subtomogram alignment and averaging were implemented using MATLAB (MathWorks) scripts derived from the TOM (37) and AV3 (38) packages as described previously (39,40).	MATLAB suggested: (MATLAB, RRID:SCR_001622)
   Using dedicated python scripts, the S trimer positions in the 3D tomograms from the subTOM procedure outlined above were converted into 2D positions and defocus values in the corresponding tilt series images, as well as Euler angles in the RELION convention.	python suggested: (IPython, RRID:SCR_001658)
   In order to deal with the CTF pre-multiplied sub-tomograms, as well as the multiplicity of each 3D voxel in the tilt series and the, a modified version of RELION was used for subtomogram refinement and classification (details to be described elsewhere).	RELION suggested: (RELION, RRID:SCR_016274)
   Movies were motion-corrected and dose-weighted using RELION’s implementation of the MotionCor2 algorithm (46).	MotionCor2 suggested: (MotionCor2, RRID:SCR_016499)
   Residues were adjusted manually in Coot 0.9 (50)	Coot suggested: (Coot, RRID:SCR_014222)
   The unmasked model-to-map FSC was calculated in PHENIX for the refined model against the full reconstruction. 3D model building of spikes on authentic virions: In order to visualize the spike protein on the authentic SARS-CoV-2 virions, the coordinates, orientations and conformational classes determined by subtomogram averaging were converted into a format compatible with Maxon Cinema 4D, and imported together with the 3D models of the different conformational states determined by subtomogram averaging.	PHENIX suggested: (Phenix, RRID:SCR_014224)
   Images of individual virions from the dataset were rendered into Adobe Photoshop to generate images for presentation.	Adobe Photoshop suggested: (Adobe Photoshop, RRID:SCR_014199)

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   Results from OddPub: Thank you for sharing your data.

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   Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

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   Results from TrialIdentifier: No clinical trial numbers were referenced.

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   Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

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   Results from JetFighter: Please consider improving the rainbow (“jet”) colormap(s) used on page 17. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.

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   Results from rtransparent:

   • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
   • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
   • No protocol registration statement was detected.

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3. 

   Version published to 10.1101/2020.06.27.174979v1 on bioRxiv

   Jun 27, 2020