Neuropilin-1 facilitates SARS-CoV-2 cell entry and infectivity

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1. 

   ScreenIT

   Mar 1, 2021

   SciScore for 10.1101/2020.06.07.137802: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	IRB: Human immunohistochemistry: All experiments with human materials were approved by the ethics committee of the University Medical Center Göttingen and were performed in accordance with the respective national, federal and institutional regulations.
   Randomization	To measure the infectivity of cultured cells, three randomly selected areas per coverslip were imaged.
   Blinding	not detected.
   Power Analysis	not detected.
   Sex as a biological variable	Adult male and female C57BL/6N mice (8 to 10 weeks of age) were taken for all experiments.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Generation of monoclonal antibodies against NRP1 b1b2: …

   SciScore for 10.1101/2020.06.07.137802: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	IRB: Human immunohistochemistry: All experiments with human materials were approved by the ethics committee of the University Medical Center Göttingen and were performed in accordance with the respective national, federal and institutional regulations.
   Randomization	To measure the infectivity of cultured cells, three randomly selected areas per coverslip were imaged.
   Blinding	not detected.
   Power Analysis	not detected.
   Sex as a biological variable	Adult male and female C57BL/6N mice (8 to 10 weeks of age) were taken for all experiments.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Generation of monoclonal antibodies against NRP1 b1b2: Female BALB/c and C57BL/6 mice, 8–9 weeks old, were immunized intraperitoneally with 17 μg of recombinant NRP1 b1b2 mixed with an equal volume of complete Freund’s adjuvant (Sigma–Aldrich Chemie, Steinheim, Germany), followed by a booster immunization four weeks later of the same dose mixed with incomplete Freund’s adjuvant (Sigma–Aldrich).	NRP1 suggested: None
   Primary antibodies were diluted in 10% blocking solution: 1:250 NRP1 (monoclonal rabbit, ab81321, Abcam); 1:1000 TuJ1 (monoclonal mouse, G712A, Promega); 1:250 NeuN (polyclonal chicken, ABN91, Milipore); 1:2000 GFP (polyclonal rabbit, A-6455, Thermo Fisher Scientific), 1:250 ColIV (polyclonal goat, 1340-01, Southern Biotech)	TuJ1 suggested: (Covance Cat# MMS-435P, RRID:AB_2313773) NeuN suggested: (Millipore Cat# ABN91, RRID:AB_11205760) GFP suggested: (Thermo Fisher Scientific Cat# PA1-86341, RRID:AB_931091)
   After three washes in PBS, the sections were incubated in secondary antibody: Alexa Fluor 488 donkey anti-mouse (R37114, Thermo Fischer Scientific);	anti-mouse ( R37114 suggested: None
   Primary antibodies were applied over night at a dilution of 1:100 for SARS-CoV S protein (monoclonal mouse, ab272420, Abcam; microwave, citric acid buffer, 10 mM, pH 6.0), 1:250 for NRP1 (monoclonal rabbit, ab81321, Abcam; microwave, Tris-EDTA, pH 8.0) and 1:150 for OLIG2 (polyclonal rabbit, 18953, IBL; microwave, Tris-EDTA, pH 8.0).	OLIG2 suggested: None
   Secondary antibodies were added as follows: biotinylated anti-mouse 1:200 (GE Healthcare RPN 1001) followed by Tyramide Super Boost with Alexa Fluor 488 1:500 (Thermo Fisher Scientific) and Alexa Fluor 555 anti rabbit 1:500 for 2 h, at room temperature.	anti suggested: None
   Either the Envision+ System for mice (Dako) and alkaline phosphatase conjugated anti-rabbit antibodies with FastBlue (Sigma Aldrich) or alkaline phosphatase coupled anti-mouse antibodies with FastBlue (Sigma Aldrich) were used for immunohistochemistry.	anti-rabbit suggested: None anti-mouse suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Cell culture: HEK 293T and Caco-2 (ATCC) cells were grown in complete growth media supplemented with 10% fetal calf serum (FCS), (pen/strep, L-Glutamine in DMEM) and passaged 1:8 (HEK-293 T) or 1:5 (Caco-2) every three	HEK 293T suggested: None Caco-2 suggested: None HEK-293 suggested: CLS Cat# 300192/p777_HEK293, RRID:CVCL_0045)
   PPC-1 human primary prostate cells were obtained from Erkki Ruoslahti’s laboratory at the Cancer Research Center,	PPC-1 suggested: ATCC Cat# HTB-190, RRID:CVCL_4778)
   Production of SARS-CoV-2 S-pseudotyped lentiviral particles: HEK-293T cells were grown in complete growth media (10% FCS, Pen/strep, L-Glutamine in DMEM) until 60 to 70% confluent in a 10 cm dish.	HEK-293T suggested: None
   Isolation of WT SARS-CoV-2 from COVID-19 patient and virus propagation: Samples were obtained under the Helsinki University Hospital laboratory research permit HUS/32/2018 § 16. 500 µl of nasopharyngeal swab in Copan UTM® Universal Transport Medium was inoculated on Calu-3 cells and incubated for 1 h in +37°C, after which the inoculum was removed and replaced with Minimum Essential Medium supplemented with 2% FBS, L-glutamine, penicillin and streptomycin.	Calu-3 suggested: KCLB Cat# 30055, RRID:CVCL_0609)
   2 were propagated in VeroE6 cells (SARSmutFC) or Caco-2 cells up to passage 7 or passage 1, respectively.	VeroE6 suggested: JCRB Cat# JCRB1819, RRID:CVCL_YQ49)
   Experimental Models: Organisms/Strains
   Sentences	Resources
   Adult male and female C57BL/6N mice (8 to 10 weeks of age) were taken for all experiments.	C57BL/6N suggested: None
   BALB/c mice were purchased from Jackson Laboratories.	BALB/c suggested: RRID:IMSR_ORNL:BALB/cRl)
   Generation of monoclonal antibodies against NRP1 b1b2: Female BALB/c and C57BL/6 mice, 8–9 weeks old, were immunized intraperitoneally with 17 μg of recombinant NRP1 b1b2 mixed with an equal volume of complete Freund’s adjuvant (Sigma–Aldrich Chemie, Steinheim, Germany), followed by a booster immunization four weeks later of the same dose mixed with incomplete Freund’s adjuvant (Sigma–Aldrich).	C57BL/6 suggested: None
   Software and Algorithms
   Sentences	Resources
   Image visualization was performed using Omero Server software 5.6	Omero Server suggested: (OME - Open Microscopy Environment, RRID:SCR_008849)
   , Omero figure 4.0.2 (https://github.com/ome/omero-figure) and InkScape 0.9234.	Omero suggested: (OMERO, RRID:SCR_002629) InkScape suggested: (Inkscape, RRID:SCR_014479)
   Image analysis: Images were analysed with CellProfiler 3.1.8 (https://cellprofiler.org/).	CellProfiler suggested: None https://cellprofiler.org/ suggested: (CellProfiler Image Analysis Software, RRID:SCR_007358)
   All other measurements were performed semiquantitatively using Fiji software 36.	Fiji suggested: (Fiji, RRID:SCR_002285)
   Primer pools targeting SARS-CoV-2 were designed using PrimalScheme tool http://primal.zibraproject.org38 and PCR was conducted using PhusionFlash PCR master mix (ThemoFisher)	ThemoFisher suggested: None
   The trimmed sequence reads were assembled to the reference sequence (NC_045512.2) using BWA-MEM algorithm implemented in SAMTools version 1.840.	BWA-MEM suggested: (Sniffles, RRID:SCR_017619) SAMTools suggested: (SAMTOOLS, RRID:SCR_002105)
   The minority variants and insertion/deletion sites were called using LoFreq* version 2.1.441. scRNA-seq analysis: Analyses of the scRNA-seq datasets including filtering, normalization and clustering were conducted using Seurat 3.142	LoFreq* suggested: None Seurat suggested: (SEURAT, RRID:SCR_007322)
   Human olfactory neuroepithelium raw data from Durante et al.22, was downloaded from Gene Expression Omnibus under accession code GSE139522.	Gene Expression Omnibus suggested: (Gene Expression Omnibus (GEO, RRID:SCR_005012)
   Statistics were performed in GraphPad Prism.	GraphPad Prism suggested: (GraphPad Prism, RRID:SCR_002798)

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   Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

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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 found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).

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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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   Read the original source
2. Version published to 10.1126/science.abd2985

   Nov 13, 2020
3. 

   ScreenIT

   Jun 10, 2020

   SciScore for 10.1101/2020.06.07.137802: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement

   Human immunohistochemistry All experiments with human materials were approved by the ethics committee of the University Medical Center Göttingen and were performed in accordance with the respective national , federal and institutional regulations .

   Randomization

   To measure the infectivity of cultured cells , three randomly selected areas per coverslip were imaged.

   Blinding

   not detected.

   Power Analysis

   not detected.

   Sex as a biological variable

   not detected.

   Cell Line Authentication

   not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Here, we found that the cellular receptor neuropilin-1 (NRP1), known to bind furin-cleaved substrates, significantly …

   SciScore for 10.1101/2020.06.07.137802: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement

   Human immunohistochemistry All experiments with human materials were approved by the ethics committee of the University Medical Center Göttingen and were performed in accordance with the respective national , federal and institutional regulations .

   Randomization

   To measure the infectivity of cultured cells , three randomly selected areas per coverslip were imaged.

   Blinding

   not detected.

   Power Analysis

   not detected.

   Sex as a biological variable

   not detected.

   Cell Line Authentication

   not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Here, we found that the cellular receptor neuropilin-1 (NRP1), known to bind furin-cleaved substrates, significantly potentiates SARS-CoV-2 infectivity, which was inhibited by a monoclonal blocking antibody against the extracellular b1b2 domain of NRP1.	neuropilin-1 suggested: None
   e , f , Representative images and quantification of HEK-293T cells expressing NRP1+ACE2+TMPRSS2 after SARS-2 pseudotype inoculation in the presence of mAb3 antibody against NRP1 ( e , mAb3 , lower panel ) or control immunoglobulin ( e , ctrl Ab , upper panel) , and in the presence of NRP1 b1b2 domain ( f , wt b1b2 , lower panel ) or the NRP1 mutant b1b2 domain ( f , mut b1b2 , upper panel) .	NRP1 mutant b1b2 domain ( f , mut b1b2 suggested: None
   Primary antibodies were diluted in 10 % blocking solution: 1:250 NRP1 ( monoclonal rabbit , ab81321 , Abcam); 1:1000 TuJ1 ( monoclonal mouse , G712A , Promega); 1:250 NeuN ( polyclonal chicken , ABN91 , Milipore); 1:2000 GFP ( polyclonal rabbit , A-6455 , Thermo Fisher Scientific) , 1:250 ColIV ( polyclonal goat , 1340-01 , Southern Biotech)	TuJ1 suggested: (Covance Cat# MMS-435P, AB_2313773) <div style="margin-bottom:8px"> <div><b>NeuN</b></div> <div>suggested: (Millipore Cat# ABN91, <a href="https://scicrunch.org/resources/Any/search?q=AB_11205760">AB_11205760</a>)</div> </div> <div style="margin-bottom:8px"> <div><b>GFP</b></div> <div>suggested: (Thermo Fisher Scientific Cat# PA1-86341, <a href="https://scicrunch.org/resources/Any/search?q=AB_931091">AB_931091</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">After three washes in PBS , the sections were incubated in secondary antibody: Alexa Fluor 488 donkey anti-mouse ( R37114 , Thermo Fischer Scientific);</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>anti-mouse ( R37114</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Primary antibodies were applied over night at a dilution of 1:100 for SARS-CoV S protein ( monoclonal mouse , ab272420 , Abcam; microwave , citric acid buffer , 10 mM , pH 6.0) , 1:250 for NRP1 ( monoclonal rabbit , ab81321 , Abcam; microwave , Tris-EDTA , pH 8.0 ) and 1:150 for OLIG2 ( polyclonal rabbit , 18953 , IBL; microwave , Tris-EDTA , pH 8.0) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>NRP1</b></div> <div>suggested: None</div> </div> <div style="margin-bottom:8px"> <div><b>OLIG2</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Secondary antibodies were added as follows: biotinylated anti-mouse 1:200 ( GE Healthcare RPN 1001 ) followed by Tyramide Super Boost with Alexa Fluor 488 1:500 ( Thermo Fisher Scientific ) and Alexa Fluor 555 anti rabbit 1:500 for 2 h , at room temperature.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>anti</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Either the Envision+ System for mice ( Dako ) and alkaline phosphatase conjugated anti-rabbit antibodies with FastBlue ( Sigma Aldrich ) or alkaline phosphatase coupled anti-mouse antibodies with FastBlue ( Sigma Aldrich ) were used for immunohistochemistry .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>anti-rabbit</b></div> <div>suggested: None</div> </div> <div style="margin-bottom:8px"> <div><b>anti-mouse</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">AgNPs ( magenta ) were visible inside the cells , counterstained with Alexa-Fluor488-phalloidin ( yellow ) and Hoechst ( cyan) . b , Representative images and quantification of NRP1-expressing HEK-293T cells treated for 30 minutes with the blocking mAb3 antibody ( lower row ) or control Ab ( upper row) . c , Representative images and quantification of NRP1-expressing HEK-293T cells incubated for 30 minutes with AgNPs , together with wild-type b1b2 domain ( wt b1b2 , lower row ) or the mutant b1b2 domain of NRP-1 ( mutant b1b2 , upper row) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>mAb3</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">NRP1 immunohistochemistry of 12 human tissues were done using NRP1 antibodies HPA030278 ( Atlas Antibodies AB ) or CAB004511 ( Santa Cruz Biotechnology ) ( brown) , and counterstained with hematoxylin ( blue) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>CAB004511</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Experimental Models: Cell Lines</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">We validated the effect of the recombinant NRP1 proteins , by showing that b1b2 , but not the mutant protein , blocked uptake of AgNP-CendR in HEK-293T cells ( Extended Data Fig .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>HEK-293T</b></div> <div>suggested: KCB Cat# KCB 200744YJ, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_0063">CVCL_0063</a></div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Confirming recent reports15 , we found that SARS-CoV-2 viruses , passaged in VeroE6 cells rapidly accumulated mutations around the furin cleavage site of the S protein .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>VeroE6</b></div> <div>suggested: JCRB Cat# JCRB1819, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_YQ49">CVCL_YQ49</a></div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Figure 2│ A blocking antibody against the b1b2 domain of NRP1 reduces infection of wild-type SARS-CoV-2 ( wt SARS-2) , but not a mutant with a deletion at the furincleavage site ( mutant SARS-2) . a , b , Representative images ( a ) and quantification ( b ) of infection assays in Caco2 cells in the presence of control mAb1 ( ctrl .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Caco2</b></div> <div>suggested: CLS Cat# 300137/p1665_CaCo-2, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_0025">CVCL_0025</a></div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Cell culture HEK 293T and Caco-2 ( ATCC ) cells were grown in complete growth media supplemented with 10 % fetal calf serum ( FCS) , ( pen/strep , L-Glutamine in DMEM ) and passaged 1:8 ( HEK-293 T ) or 1:5 ( Caco-2 ) every three days</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>HEK 293T</b></div> <div>suggested: None</div> </div> <div style="margin-bottom:8px"> <div><b>HEK-293</b></div> <div>suggested: CLS Cat# 300192/p777_HEK293, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_0045">CVCL_0045</a></div> </div> <div style="margin-bottom:8px"> <div><b>Caco-2</b></div> <div>suggested: CLS Cat# 300137/p1665_CaCo-2, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_0025">CVCL_0025</a></div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">For confocal imaging of NRP-1 expressing PPC-1 cells , 30,000 cells/well were cultured at 37°C in 5 % CO2 as a monolayer on coverslips ( d = 12 mm; Paul Marienfeld GmbH & Co .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>PPC-1</b></div> <div>suggested: ATCC Cat# HTB-190, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_4778">CVCL_4778</a></div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Isolation of WT SARS-CoV-2 from COVID-19 patient and virus propagation Samples were obtained under the Helsinki University Hospital laboratory research permit HUS/32/2018 § 16 . 500 µl of nasopharyngeal swab in Copan UTM® Universal Transport Medium was inoculated on Calu-3 cells and incubated for 1 h in +37°C , after which the inoculum was removed and replaced with Minimum Essential Medium supplemented with 2 % FBS , L-glutamine , penicillin and streptomycin .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Calu-3</b></div> <div>suggested: BCRJ Cat# 0264, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_0609">CVCL_0609</a></div> </div> </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Software and Algorithms</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Mercer , J . , Schelhaas , M. & Helenius , A . Virus entry by endocytosis .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Mercer</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Image visualization was performed using Omero Server software 5.6</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Omero Server</b></div> <div>suggested: (OME - Open Microscopy Environment, <a href="https://scicrunch.org/resources/Any/search?q=SCR_008849">SCR_008849</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">, Omero figure 4.0.2 ( https://github.com/ome/omero-figure ) and InkScape 0.9234 .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Omero</b></div> <div>suggested: (OMERO, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002629">SCR_002629</a>)</div> </div> <div style="margin-bottom:8px"> <div><b>InkScape</b></div> <div>suggested: (Inkscape, <a href="https://scicrunch.org/resources/Any/search?q=SCR_014479">SCR_014479</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Image analysis Images were analysed with CellProfiler 3.1.8 ( https://cellprofiler.org/).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>CellProfiler</b></div> <div>suggested: None</div> </div> <div style="margin-bottom:8px"> <div><b>https://cellprofiler.org/</b></div> <div>suggested: (CellProfiler Image Analysis Software, <a href="https://scicrunch.org/resources/Any/search?q=SCR_007358">SCR_007358</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">All other measurements were performed semiquantitatively using Fiji software 36 .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Fiji</b></div> <div>suggested: (Fiji, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002285">SCR_002285</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Primer pools targeting SARS-CoV-2 were designed using PrimalScheme tool http://primal.zibraproject.org 38 and PCR was conducted using PhusionFlash PCR master mix ( ThemoFisher)</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>ThemoFisher</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The trimmed sequence reads were assembled to the reference sequence ( NC_045512.2 ) using BWA-MEM algorithm implemented in SAMTools version 1.840 .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>BWA-MEM</b></div> <div>suggested: (Sniffles, <a href="https://scicrunch.org/resources/Any/search?q=SCR_017619">SCR_017619</a>)</div> </div> <div style="margin-bottom:8px"> <div><b>SAMTools</b></div> <div>suggested: (Samtools, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002105">SCR_002105</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The minority variants and insertion/deletion sites were called using LoFreq* version 2.1.441 . scRNA-seq analysis Analyses of the scRNA-seq datasets including filtering , normalization and clustering were conducted using Seurat 3.142</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>LoFreq*</b></div> <div>suggested: None</div> </div> <div style="margin-bottom:8px"> <div><b>Seurat</b></div> <div>suggested: (SEURAT, <a href="https://scicrunch.org/resources/Any/search?q=SCR_007322">SCR_007322</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Human olfactory neuroepithelium raw data from Durante et al.22 , was downloaded from Gene Expression Omnibus under accession code GSE139522 .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Gene Expression Omnibus</b></div> <div>suggested: (Gene Expression Omnibus (GEO), <a href="https://scicrunch.org/resources/Any/search?q=SCR_005012">SCR_005012</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Statistics were performed in GraphPad Prism .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>GraphPad Prism</b></div> <div>suggested: (GraphPad Prism, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002798">SCR_002798</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">ImageJ2: ImageJ for the next generation of scientific image data .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>ImageJ</b></div> <div>suggested: (ImageJ, <a href="https://scicrunch.org/resources/Any/search?q=SCR_003070">SCR_003070</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Multiplex PCR method for MinION and Illumina sequencing of Zika and other virus genomes directly from clinical samples .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>MinION</b></div> <div>suggested: (MinION, <a href="https://scicrunch.org/resources/Any/search?q=SCR_017985">SCR_017985</a>)</div> </div> </td></tr></table> Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015). Results from OddPub: We did not find a statement about open data. We also did not find a statement about open code. Researchers are encouraged to share open data when possible (see Nature blog). About SciScore SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore is not a substitute for expert review. SciScore checks for the presence and correctness of RRIDs (research resource identifiers) in the manuscript, and detects sentences that appear to be missing RRIDs. SciScore also checks to make sure that rigor criteria are addressed by authors. It does this by detecting sentences that discuss criteria such as blinding or power analysis. SciScore does not guarantee that the rigor criteria that it detects are appropriate for the particular study. Instead it assists authors, editors, and reviewers by drawing attention to sections of the manuscript that contain or should contain various rigor criteria and key resources. For details on the results shown here, including references cited, please follow this link.

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   Version published to 10.1101/2020.06.07.137802 on bioRxiv

   Jun 7, 2020