How Antioxidants, Osmoregulation, Genes and Metabolites Regulate the Late Seeding Tolerance of Rapeseeds (Brassica napus L.) during Wintering

Round 1

Reviewer 1 Report

The article investigates the impact of delayed seeding dates on rapeseed growth and adaptation under a rice-rape-rotation system. The study employs five different seeding dates ranging from October 20th to November 30th and explores the differences in growth and stress tolerance between late-seeding-tolerant (LST) and late-seeding-sensitive (LSS) rapeseed varieties through a field experiment.

Overall, the study presents novel insights into the effects of delayed seeding on rapeseed performance. The manuscript is well organized and only few points need to be addressed before the publication in the journal.

Details:

-          Introduction: Some sentences are quite lengthy and complex. Consider simplifying them for clarity.

Line 39 Add the recent reference: Antioxidants 2023, 12, 1621. https://doi.org/10.3390/antiox12081621

-          Results: The results section presents interesting findings related to rapeseed growth and agronomic traits under varying seeding dates, emphasizing the differences between late-seeding-tolerant (LST) and late-seeding-sensitive (LSS) rapeseed varieties. All the Figures are difficult to read, increase the font size to make them clearer

-          Conclusions could be enhanced by providing a more detailed and comprehensive discussion of the relevance and practical implications of the results.

Author Response

Reviewer1: The article investigates the impact of delayed seeding dates on rapeseed growth and adaptation under a rice-rape-rotation system. The study employs five different seeding dates ranging from October 20th to November 30th and explores the differences in growth and stress tolerance between late-seeding-tolerant (LST) and late-seeding-sensitive (LSS) rapeseed varieties through a field experiment. Overall, the study presents novel insights into the effects of delayed seeding on rapeseed performance. The manuscript is well organized and only few points need to be addressed before the publication in the journal. Author’s answer: Thanks a lot for your appreciation to our findings and kindly proposals for the improvements of our manuscript, thank you very much. And the manuscript was revised accordingly. Details: - Introduction: Some sentences are quite lengthy and complex. Consider simplifying them for clarity. Author’s answer: Thank you for your comments, several lengthy and complex sentences were rewritten for clarity, for example in Line 44-45, 47-50, 54-58, 70-72, 86-88, 93-100, and so on. Line 39 Add the recent reference: Antioxidants 2023, 12, 1621. https://doi.org/10.3390/antiox12081621. Author’s answer: Thank you for this suggestion, this reference was added in Line 39. - Results: The results section presents interesting findings related to rapeseed growth and agronomic traits under varying seeding dates, emphasizing the differences between late-seeding-tolerant (LST) and late-seeding-sensitive (LSS) rapeseed varieties. All the Figures are difficult to read, increase the font size to make them clearer Author’s answer: Thank you very much for this key point, we have regulated the font size accordingly for example the heatmap in figure 1, figure 2 and figure 5, the font size of figure 2 (A-D) and figure 4 (A-C) were amplified, and few of the figures were remained because of the combination of numerous subgraphs, and we will notice it to make our manuscript clearer in future, thank you. - Conclusions could be enhanced by providing a more detailed and comprehensive discussion of the relevance and practical implications of the results. Author’s answer: Thank you for your suggestion, the detailed discussion was added in subsection 4.3 (Line 468-489). The practical implications of the results were added in Conclusion part (Line 503-508).

Author Response File: Author Response.pdf

Reviewer 2 Report

The manuscript by Hao et al. described a field trial study on rapeseed seeding in combination with imaging-based phenotyping, enzyme activity assays and Omics analyses. Overall, it presents a well-designed research with a clear goal to evaluate how rapeseeds respond to delayed growth environments. My major concern lies mostly in the writing aspect, as there is significant need for clarity and improved context and figures throughout the manuscript.

Line 52-54 The statement about temperature change here is not clear. Please be more specific.

Line 60-64 Please follow proper English grammar to clearly structure the sentence like this one throughout the manuscript.

Line 112 What is “long-term rice-rapeseed rotation”?

Line 193-204 Please cite each panel of Figure 1 at right place in this section.

Figure 1 Please briefly describe D) and E) in the figure legend. For example, what do the colored bars represent?

Line 190 Remove “3.1. Subsection”.

Line 212 and 216 It is vague to say that agronomic traits are higher or lower. Please be more specific.

Line 252 “murderer” may be not the correct word here.

Figure 5 For all figures, it is better to display those four genotypes with their designated groups, i. e. the LST or LSS group. It will help for reading the text and connecting that to the figures.

Section 3.8 Please refrain from using subjective words, such as “complicated”, “easier” when interpreting omics data. Less number of DEGs identified doesn’t necessarily mean the pathway is easier.

Many descriptive sentences in the discussion part are more like the background information (Line 472-483) or redundancy in presenting the results (Line 491-500). Please focus on discussing how the results align with or deviate from previous studies and how they advances our understudying of this topic.

Line 520-521 It is an overstatement. The “adaption mechanisms” are not well revealed in this study.

There is significant need for clarity and improved context throughout the manuscript.

Author Response

Reviewer2:

The manuscript by Hao et al. described a field trial study on rapeseed seeding in combination with imaging-based phenotyping, enzyme activity assays and Omics analyses. Overall, it presents a well-designed research with a clear goal to evaluate how rapeseeds respond to delayed growth environments. My major concern lies mostly in the writing aspect, as there is significant need for clarity and improved context and figures throughout the manuscript.

Author’s answer: Thanks a lot for your recognition to our work, and we are extremely grateful to your carefully review, positive comments and constructive suggestions. The manuscript was revised carefully according to your suggestions. Furthermore, extensive English editing was made throughout the text according to your suggestion.

Line 52-54 The statement about temperature change here is not clear. Please be more specific.

Author’s answer: Thank you for this suggestion, we have corrected it to make it specifically.

Line 60-64 Please follow proper English grammar to clearly structure the sentence like this one throughout the manuscript.

Author’s answer: Thank you for this suggestion, we have corrected some of the sentence structure, such as Line 48-50, 56-68, 77, 86-88, 93-99, 400, 414-417, 441-444, 447-448, and so on.

Line 112 What is “long-term rice-rapeseed rotation”?

Author’s answer: This sentence was used to explain the background of cropping pattern at this district, and rice-rapeseed rotation were practiced for a long time before. We have corrected the description in Line 111 to avoid ambiguity.

Line 193-204 Please cite each panel of Figure 1 at right place in this section.

Author’s answer: Thank you for this suggestion, we have added some citation of figures to make the description more specifically.

Figure 1 Please briefly describe D) and E) in the figure legend. For example, what do the colored bars represent?

Author’s answer: Thanks for your suggestion, and we have added the description for this subfigure.

Line 190 Remove “3.1. Subsection”.

Author’s answer: Thank you for pointing out this mistake, and we have removed it.

Line 212 and 216 It is vague to say that agronomic traits are higher or lower. Please be more specific.

Author’s answer: Thank you for this comment, and we have corrected the description accordingly.

Line 252 “murderer” may be not the correct word here.

Author’s answer: Thank you for this comment, we have corrected it to ‘stressor’.

Figure 5 For all figures, it is better to display those four genotypes with their designated groups, i. e. the LST or LSS group. It will help for reading the text and connecting that to the figures.

Author’s answer: Thank you for this suggestion and we have added their designated groups (LST, LSS) beside the legend in Figure 5A and 5H.

Section 3.8 Please refrain from using subjective words, such as “complicated”, “easier” when interpreting omics data. Less number of DEGs identified doesn’t necessarily mean the pathway is easier.

Author’s answer: Thanks for your comments, these words were removed to make the description more scientific.

Many descriptive sentences in the discussion part are more like the background information (Line 472-483) or redundancy in presenting the results (Line 491-500). Please focus on discussing how the results align with or deviate from previous studies and how they advances our understudying of this topic.

Author’s answer: Thanks a lot for these comments, we have modified the descriptive sentences in subsection 4.2 to Line 441-444 and 447-448, and the redundancy description of results in subsection 4.3 was removed. At the same time, some discussions were added and rearranged in subsection 4.3 (Line 468-489) to improve the discussion.

Line 520-521 It is an overstatement. The “adaption mechanisms” are not well revealed in this study.

Author’s answer: Thanks for this comment, we have removed this to avoid overstatement.

 

Author Response File: Author Response.pdf

Reviewer 3 Report

In this study, the impact of delayed seeding dates on rapeseed cultivation under a rice-rape-rotation system was explored. Five distinct seeding dates ranging from late October to late November were examined, revealing significant differences between two categories of rapeseed varieties: late-seeding-tolerant (LST) and late-seeding-sensitive (LSS). As the seeding delay increased, growth was noticeably inhibited, and the yield foldchange (LST/LSS) rose from 1.50 at T1 to 2.64 at T5. Visual and hyperspectral imaging, along with data from unmanned aerial vehicles, demonstrated that LST varieties exhibited superior appearances compared to LSS varieties. Furthermore, various stress assessments revealed that LSS varieties suffered more damage. Enzymatic and osmoregulatory analyses indicated a decrease in antioxidant enzymes and osmoregulation substances with delayed seeding, while LST varieties maintained higher levels. Transcriptome and metabolome comparisons identified numerous pathways, genes, and metabolites involved in late-seeding-tolerance regulation. This study provides valuable insights into the intricate mechanisms governing rapeseed adaptation to late-seeding conditions, promising potential advancements in rapeseed genetics. Nonetheless, there are some issues that need to be clarified before publication:

-          Keywords should be arranged alphabetically.

-          Throughout the text, the authors are incorrectly using the term ‘variety’ instead of ‘cultivar’.

-          Please follow the MDPI formatting style in the references (in the text and Reference list).

-          Add the geographical location of the experimental site.

-          Units should be given as product, e.g. mg*L-1 (not mg/L).

-      Do not repeat Materials and Methods in the Results section (e.g. in lines 276-277).

-          Avoid using references in the Results chapter. Some parts of the Results are in fact a Discussion (e.g. page 12).

After making all the necessary changes, the article can be accepted for publication.

Minor grammar corrections are needed.

Author Response

Reviewer3:

In this study, the impact of delayed seeding dates on rapeseed cultivation under a rice-rape-rotation system was explored. Five distinct seeding dates ranging from late October to late November were examined, revealing significant differences between two categories of rapeseed varieties: late-seeding-tolerant (LST) and late-seeding-sensitive (LSS). As the seeding delay increased, growth was noticeably inhibited, and the yield foldchange (LST/LSS) rose from 1.50 at T1 to 2.64 at T5. Visual and hyperspectral imaging, along with data from unmanned aerial vehicles, demonstrated that LST varieties exhibited superior appearances compared to LSS varieties. Furthermore, various stress assessments revealed that LSS varieties suffered more damage. Enzymatic and osmoregulatory analyses indicated a decrease in antioxidant enzymes and osmoregulation substances with delayed seeding, while LST varieties maintained higher levels. Transcriptome and metabolome comparisons identified numerous pathways, genes, and metabolites involved in late-seeding-tolerance regulation. This study provides valuable insights into the intricate mechanisms governing rapeseed adaptation to late-seeding conditions, promising potential advancements in rapeseed genetics. Nonetheless, there are some issues that need to be clarified before publication:

Author’s answer: Thanks a lot for your high recognition to our work, and great appreciation to your kindly and carefully comments on our manuscript, which were very helpful for its improvement.

-          Keywords should be arranged alphabetically.

Author’s answer: Thank you for your suggestion, we have rearranged the keywords, and we will also pay attention to avoid this problem in the future.

-          Throughout the text, the authors are incorrectly using the term ‘variety’ instead of ‘cultivar’.

Author’s answer: Thank you for pointing out the mistakes in the manuscript, we have corrected ‘Variety’ to ‘Cultivar’ accordingly, for example, in Line 16, 27, 115, 117, 199-200, 209, 214, 225, 227-235, 243-247, 268, 280, 293-303, 324, 364, 374, 432-433, and so on.

-          Please follow the MDPI formatting style in the references (in the text and Reference list).

Author’s answer: Thank you for this suggestion, the formatting of references was changed according MDPI formatting style.

-          Add the geographical location of the experimental site.

Author’s answer: Thanks for your suggestion, the geographical location of the experiment site was added in subsection 2.1.1.

-          Units should be given as product, e.g. mg*L-1 (not mg/L).

Author’s answer: Thank you a lot for this point, we have corrected them accordingly in Line 222-223 and Line 286-288.

-      Do not repeat Materials and Methods in the Results section (e.g. in lines 276-277).

Author’s answer: Thank you very much for this comment, we have removed the first sentence which repeat Materials and Methods in the first paragraph of subsection 3.4 and subsection 3.8.

-          Avoid using references in the Results chapter.

Author’s answer: Thank you for this suggestion, these sentences with references were removed out in subsection 3.3, subsection 3.5 and subsection 3.8.

Some parts of the Results are in fact a Discussion (e.g. page 12).

Author’s answer: Thank you for this suggestion, the discussion in subsection 3.8 were moved to discussion part 4.3, which will enhance the discussion of our manuscript and avoid discussion in Results part, thank you for this suggestion.

After making all the necessary changes, the article can be accepted for publication.

 

Author Response File: Author Response.pdf

Round 2

Reviewer 2 Report

I saw some desired clarifications have been made. However, the overall quality of presentation still needs substantial improvement. For example, terms such as "better appearance" mentioned in the abstract are way too general; What does that mean? Please be more specific and present results in a more scientific way. Some sentences such as "The air temperature decreased very quickly from November to December from 30℃ maximum to -2℃ minimum, which will amplify the influences of severe environment conditions on rapeseed production" are still very confusing. It is hard to comprehend what the authors' intension to describe. Are they the temperature records of field locations?  

 

Needs improvement.

Author Response

Reviewer1: I saw some desired clarifications have been made. However, the overall quality of presentation still needs substantial improvement. Author’s answer: thank you very much for your appreciation to our modification on our manuscript. The overall quality of presentation and writing was improved according to the proposition of editors at https://www.mdpi.com/authors/english. The following questions were modified accordingly. For example, terms such as "better appearance" mentioned in the abstract are way too general; What does that mean? Please be more specific and present results in a more scientific way. Author’s answer: thank you very much for this suggestion, we have corrected this description of ‘better appearance’ to ‘higher plant coverage’, which were more specific and scientific. Some sentences such as "The air temperature decreased very quickly from November to December from 30℃ maximum to -2℃ minimum, which will amplify the influences of severe environment conditions on rapeseed production" are still very confusing. It is hard to comprehend what the authors' intension to describe. Are they the temperature records of field locations? Author’s answer: thank you for your comments, we aimed to illustrate the sharp decreasing of air temperature as the delay of seeding dates, while such low temperature was not conducive to rapeseed growth. We have changed this sentence to ‘Meanwhile, the maximum air temperature decreases very quickly from near 30℃ in November to even below 5℃in December, which greatly inhibit the production of rapeseed’ to avoid confusing’. The temperatures were the records of this field locations. Thank you again for your careful review, which help us a lot for the improvement on our manuscript. Thank you very much.

Author Response File: Author Response.docx

Round 3

Reviewer 2 Report

Most of the necessary clarifications have been provided. TI have no addition comments. I recommend its publication.

The writing is improved and now the text is easy to read and comprehend.