PONE-D-20-23256

Comparative proteomics analysis reveals the molecular mechanism of enhanced cold tolerance through ROS scavenging in winter rapeseed (Brassica napus L.)

PLOS ONE

Dear Dr. Liu,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

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We look forward to receiving your revised manuscript.

Kind regards,

Maoteng Li

Academic Editor

PLOS ONE

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2.Thank you for stating the following in the Acknowledgments Section of your manuscript:

[This work was supported by the National Natural Science

Foundation of China (31660404), National Key Basic Research and Development

Program (2018YFD0100500), University of Gansu Province Scientific Research

Achievement Transformation and Cultivation Project (2018D-13), Special Fund for

the Construction of Modern Agricultural Industrial Technology System of Gansu

Province (17ZD2NA016-4).]

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

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2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: No

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3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

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4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: No

Reviewer #2: No

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5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: In this manuscript, the authors reported two winter rapeseed cultivars, cold tolerant and cold sensitive, were used to reveal the morphological, physiological, and proteomic characteristics in leaves of plants after treatment at different temperature points. The candidate pathways of DAPs involved were analyzed and ROS scavenging pathway may play function in rapeseed cold tolerance. The paper is poorly displayed and following items should be modified.

1. The format of references is incorrect, please correct it.

2. The description of Result 2.2, “…identified based on the criteria of p < 0.005 for the adjusted p-value…”, is confused.

3. An obvious descriptive error existed in Methods 4.1, “…store at -80 reaching concentrations thaEach sample….”.

4. Abstract part, the first line, it is should be “cultivars” not “culvers”. It is recommended to find native English speaker to modify the language of the paper.

4. It is confused to understand if the proteomics performed with biological repetition?

Reviewer #2: In this work, the authors performed comparative proteomics analysis reveals the molecular mechanism of enhanced cold tolerance through ROS scavenging in winter rapeseed (Brassica napus L.). Although this work is interesting, the manuscript is not well organized, and the data are not well presented. Therefore, the paper needs to be thoroughly revised.

1. The authors must carefully proof-read the manuscript to minimize typographical, grammatical, and bibliographic errors. Some typographical errors, some nonsensical or confusing sentences should be corrected.

2. In Abstract, “Two winter rapeseed culvers, “NF” (cold tolerant) and “NS” (cold sensitive), were used to reveal the morphological, physiological, and proteomic characteristics……”. What does ‘culvers’ mean? ‘culvers’ or ‘cultivars’? Please check the spelling.

3. In Abstract, authors refer to two winter rapeseed culvers “NF” and “NS”, among, “NF” represent cold-tolerant cultivar and “NS” represent cold-sensitive cultivar. However, in Introduction and Methods parts, authors refer to “strong cold-tolerant variety “NS” and '17NTS57' (abbreviated as “NS”, cold-tolerant…) and 'NQF24' (abbreviated as “NF”, cold-sensitive…) winter rapeseed (Brassica napus L.). These seem to be particularly controversial. Please confirm and explain why?

4. In Plant materials, cold stress treatment and physiological part, authors say “the cold-tolerant variety ‘17NTS57’ beyond 90% survival rate under −26 °C of extreme low temperature during overwintering, please provide data supporting the result or cite the related references.

5. In results part, ‘(figure2B )’ should put it after the sentence ‘By comparing all of the DAPs between the two varieties at three treated time points, the Venn diagram shows the number of more- or less-abundant DAPs as well as specific and commonly abundant DAPs in the different groups.’

6. In general, the technical approaches of RT-qPCR seem fine. However, statistics for figure 6 are not given. Also, statistics for DAGs (real-time quantitative RT-PCR) abundance changes (eg p-values) are not given (not in method and not in legend). What method were used? ANOVA? One or two way? What was the level of significance?

7. Authors selected 23 DAPs for detecting the mRNA levels using qRT-PCR. However, the protein names are not given in figure 6 or in Result part. The protein names are replaced by Numbers 1, 2, 3, 4, 5……

8. In Result part, no any information and sentences related to figure 7. Please add …..!

9. The results lack systematic in-depth analyses. Authors think cold-tolerant rapeseed enhance cold tolerance through ROS scavenging, but no related candidate proteins were in-depth analyzed and studied.

10. Figure quality is too low and the legends need to be reorganized and rephrased.

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6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

PONE-D-20-23256R1

Comparative proteomics analysis reveals the molecular mechanism of enhanced cold tolerance through ROS scavenging in winter rapeseed (Brassica napus L.)

PLOS ONE

Dear Dr. Liu,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

Please submit your revised manuscript by Dec 25 2020 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

• A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
• A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
• An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols

We look forward to receiving your revised manuscript.

Kind regards,

Maoteng Li

Academic Editor

PLOS ONE

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

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2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The authors modified all the concerns we raised, I think it can be accepted with a minor revision.

1. Please write the full name for the abbreviation of AVG when it first appeared.

2. In methods part, the real-time quantitative PCR, “The Actin gene was used as an internal reference…..”, the accession number for the actin gene used should be listed in the paper.

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7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

Comparative proteomics analysis reveals the molecular mechanism of enhanced cold tolerance through ROS scavenging in winter rapeseed (Brassica napus L.)

PONE-D-20-23256R2

Dear Dr. Liu,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

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Kind regards,

Maoteng Li

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments: