Rescue of SARS-CoV-2 from a single bacterial artificial chromosome

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   ScreenIT

   Jul 25, 2020

   SciScore for 10.1101/2020.07.22.216358: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	IACUC: Biosafety (IBC) and Animal Care and Use (IACUC) committees.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Sex as a biological variable	Pathogenicity studies in golden Syrian hamsters: Twelve-week-old female golden Syrian hamsters were purchased from Charles River and maintained in the animal facility at Texas Biomed under specific pathogen-free conditions.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Cells were incubated overnight with 1 μg/ml of a SARS-CoV cross-reactive N monoclonal antibody 1C7 at 4°C, washed with PBS, and stained with a FITC-labeled goat anti-mouse IgG (1:200).	anti-mouse IgG

   SciScore for 10.1101/2020.07.22.216358: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	IACUC: Biosafety (IBC) and Animal Care and Use (IACUC) committees.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Sex as a biological variable	Pathogenicity studies in golden Syrian hamsters: Twelve-week-old female golden Syrian hamsters were purchased from Charles River and maintained in the animal facility at Texas Biomed under specific pathogen-free conditions.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Cells were incubated overnight with 1 μg/ml of a SARS-CoV cross-reactive N monoclonal antibody 1C7 at 4°C, washed with PBS, and stained with a FITC-labeled goat anti-mouse IgG (1:200).	anti-mouse IgG suggested: None
   Then, cells were blocked with 2.5% bovine serum albumin (BSA) in PBS for 1 h at 37 °C, followed by incubation with 1 µg/ml of the anti-N SARS-CoV monoclonal antibody 1C7 diluted in 1% BSA for 1 h at 37 °C.	anti-N SARS-CoV suggested: (MyBioSource Cat# MBS432038, RRID:AB_2184133)
   Experimental Models: Cell Lines
   Sentences	Resources
   Briefly, confluent monolayers of Vero E6 cells (106 cells/well, 6-well plates, triplicates) were transfected, using LPF2000, with 4.0 μg/well of SARS-CoV-2 BAC, or empty BAC as internal control.	Vero E6 suggested: RRID:CVCL_XD71)
   Software and Algorithms
   Sentences	Resources
   Raw reads were quality filtered using Trimmomatic v0.39 (32) and mapped to a SARS-CoV-2 reference genome (Genbank Accession No. MN985325) with Bowtie2 v2.4.1 (33)	Trimmomatic suggested: (Trimmomatic, RRID:SCR_011848) Bowtie2 suggested: (Bowtie 2, RRID:SCR_016368)
   We genotyped each sample for low frequency variants with LoFreq* v2.1.3.1 (35) and filtered sites with less than 100x read depth or minor allele frequencies less than 1%.	LoFreq* suggested: None
   Finally, we used SnpEff v4.3t (36) to identify the impact of potential variants on the protein coding regions in the SARS-CoV-2 reference genome.	SnpEff suggested: (SnpEff, RRID:SCR_005191)
   After incubation with the primary antibody, cells were washed three times with PBS, counterstained with the Vectastain ABC kit, and developed using the DAB Peroxidase Substrate kit (Vector Laboratory, Inc, CA, USA) according to the manufacturers’ instructions.	Vector Laboratory suggested: None
   Evaluation of lung pathological lesions: Macroscopic pathology scoring was evaluated using ImageJ software to determine the percent of the total surface area of the lung (dorsal and ventral view) affected by consolidation, congestion, and atelectasis, as previously described (38).	ImageJ suggested: (ImageJ, RRID:SCR_003070)

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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: We did not find any issues relating to colormaps.

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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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   Version published to 10.1101/2020.07.22.216358 on bioRxiv

   Jul 22, 2020