Variability in Leaf Color Induced by Chlorophyll Deficiency: Transcriptional Changes in Bamboo Leaves

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript by Pengkai et al. uses bioinformatic and biochemical approaches to investigate transcriptional changes in bamboo leaves and their correlation with chlorophyll deficiency. The authors have generated a solid amount of data and did bioinformatic analysis. Here, I include some comments and suggestions.

 

1. In Methods part.

Line 103: typo “Gigments”.

 

2. In Results part.

Figure 1: A, it would be better to label the leaf types as “G”, “W”, and “GW” in figure and indicate the leaf stage in legends.

Line 166: “(Figure 1E; Figure 1G)”, no Fig. 1G in Fig. 1.

Figure 2: how to interpret the left panel in A and B. Wat means the density of red or blue circles? Please include detailed explanations in legends.

Figure 3: Please a general title before individual sub-figures. Details in legends, e.g. which statistical analysis, …

 

3. In Discussion part.

Figure 5: Extract symbols in “Chlorophyll P680” and “Chlorophyll P700”.  

Author Response

Thank you for your thorough review and valuable feedback. We greatly appreciate your insights, which have helped us address overlooked details and enhance the manuscript's completeness and readability.

In response to your suggestions:

For Figure 1, we've added leaf type labels "G", "W", and "GW" next to the sample photos in panel A and clarified the leaf stages in the legend for better understanding of the experimental design and results.

Regarding the reference error in Figure 1, we apologize for the confusion caused by the mention of a non-existent Figure 1G and have removed this reference. We've also introduced a new table listing key differentially expressed genes related to chlorophyll deficiency, along with their expression fold changes and confidence levels, for a clearer presentation of these findings.

For the interpretation of Figure 2, we realized the co-enrichment analysis visualization in panels A and B might not have been intuitive. Following your advice, we've detailed the significance of the point density and color differences in the legend to aid reader comprehension of these complex data.

In Figure 3, we added a general title for all sub-figures and expanded the legend to include the statistical analysis methods used and the meaning of letter markings, enhancing the clarity and transparency of the information presented.

For the symbol clarification in Figure 5, symbols for "Chlorophyll P680" and "Chlorophyll P700" have been clearly marked to ensure scientific accuracy and consistency.

In the Results section, we rearranged segments for logical flow and added a table detailing key gene expression changes, increasing result transparency (L. 211).

Additionally, we've focused the discussion by removing redundant descriptions of chlorophyll's role and added tables to support enzyme-related discussions (L.295, 357).

We hope these adjustments meet your expectations and further improve the manuscript's quality. Thank you again for your detailed review and constructive suggestions. We look forward to any further feedback you may have.

Reviewer 2 Report

Comments and Suggestions for Authors

Need minor revision

Comments for author File: Comments.pdf

Comments on the Quality of English Language

Author Response

Thank you for your thorough review and insightful comments, which have greatly contributed to the refinement of our manuscript.

In response to your suggestions, we have made several key revisions:

We clarified the research gaps and objectives in the Introduction (L. 46-59), and provided more context on why Bambusa multiplex f. silverstripe was selected for this study, including its ecological significance and unique role in leaf color diversity research (L. 36-45).

The Materials and Methods section has been reorganized for better clarity and ease of replication. We added detailed information on plant materials, growth conditions, and sample processing (L. 62-77). It's important to note that liquid nitrogen was used solely for rapid freezing of samples to prevent RNA degradation and is unrelated to soil nitrogen content. Given the focus on physiological and transcriptional changes associated with chlorophyll deficiency, soil nitrogen levels were not measured. However, your point on soil elements potentially affecting leaf color is well-taken, and we've suggested environmental impacts on leaf color as a direction for future research (L. 373-377).

We've incorporated necessary citations and converted passive voice to active voice throughout the manuscript, particularly in the Methods and Results sections, to enhance readability and directness (e.g., L. 66, 90, 105, 116).

In the Results section, we rearranged content for coherence and added a table detailing key gene expression changes, improving result transparency (L. 211).

For the Discussion, we condensed the description of results and removed repetitive discussions on chlorophyll's role within organisms, focusing more on key findings. Additionally, we added two tables to detail genes and their enzymes involved in ALA biosynthesis and sugar metabolism, supporting our discussion on enzymatic roles (L. 295, 357).

We hope these revisions address your concerns and enhance the manuscript. We appreciate your constructive feedback and look forward to any further suggestions.

Reviewer 3 Report

Comments and Suggestions for Authors

The study aims to shed light on the complex gene interactions and regulatory networks underlying leaf colour diversity in Bambusa multiplex f. Silverstripe, providing insights for future research and plant breeding. The paper is well written. Only minor adjustment is required.

Line 22; The introduction doesn't explicitly state the research gap or the specific objectives of the study. Including these elements would provide clarity on what the researchers aim to address or discover.

The text tends to use passive voice frequently, which can make the writing less engaging. Active voice could be employed to enhance clarity and directness.

While the introduction mentions the plant, it lacks sufficient context on why Bambusa multiplex f. silverstripe is chosen for the study. Providing more information on its ecological importance, uniqueness, or relevance to the research question would strengthen the introduction.

There is some repetition of information, especially regarding the involvement of genetic factors in chlorophyll synthesis. Streamlining the content and avoiding redundancy would enhance the clarity and focus of the introduction

Line 41-42; There are instances of awkward phrasing, such as "laying the foundation for future research and applications." It might be helpful to rephrase for smoother flow and clarity.

Line 118; The text contains several abbreviations, without initial explanations. It's essential to introduce and define these abbreviations upon first use to enhance reader understanding.

 

Line 378; Outline specific areas or questions that future research should address, considering the gaps identified in the current study. This helps guide researchers interested in continuing the work.

Comments on the Quality of English Language

Mino correction

Author Response

Thank you for your thorough review and valuable feedback. Your suggestions have led us to refine the details of our study, enhancing its integrity and readability. In response, we've made the following adjustments:

We've clarified the research gap and objectives in the introduction, providing a clear statement of the study's aims and expanding on the choice of Bambusa multiplex f. silverstripe, including its ecological significance and unique role in leaf color diversity research (L. 36-45).

We've revised the manuscript for more active voice, particularly in the methods and results sections, to improve clarity and engagement (e.g., L.66, 90, 105, 116).

In the results section, we've defined all abbreviations at their first occurrence and added a table detailing key genes and their expression changes for greater transparency (L. 168, 182, 211).

Additionally, we've focused the discussion by removing redundant descriptions of chlorophyll's role and added tables to support enzyme-related discussions (L.295, 357).

We've discussed future research directions in the conclusion, emphasizing the exploration of Silverstripe's leaf color variation in relation to environmental factors and the importance of population genetic analysis (L. 365-372).

We believe these revisions address your concerns and enhance the manuscript's quality. We look forward to any further feedback.

Reviewer 4 Report

Comments and Suggestions for Authors

 

Abstract 

Please add the important finding of this study and be more focused 

Introduction 

1-The introduction provides a brief overview of the research topic. However, it would be beneficial to expand upon the introduction by providing more background information about the importance and relevance of the research topic. Additionally, it would be helpful to include a clear statement of the research objectives or questions that the study aims to address.

2-Line 41 please add more references which used RNA-seq related to your study

Materials 

Please addd the name of genotype or variety used and add new section about growth conditions.

I think that the material section would benefit from being reordered as follows;

1-plant materials and growth conditions

2-Measurement of Photosynthetic Gigments and Nutrient Concentration

3-Transcriptome Sequencing

 

Results 

Directly state the results with significant findings. Make the results section concise and specific.
Please reorder the results section to be as follows;

1-Photosynthetic Pigments and Nutrient Concentrations of Three Color Types

2-Summary of Sequencing Data and Assembly

Line 121-127: rephrase and be concentrated

Line 166 Please add new table of the important identified genes and it’s fold change related to 

 

chlorophyll deficiency

Discussion

 

please reduce the detailed explanation of results in the discussion. Focus on highlighting one or two key findings and how they address your research objectives.

Conclusion

The conclusion is excessively long.

 

Comments on the Quality of English Language

Minor editing required 

Author Response

Thank you for your thorough review and valuable feedback. We've made the following revisions to enhance our manuscript's clarity and depth:

In the Introduction, we added a clear statement of research gaps and specific objectives (L. 36-45), enriching the context around why Bambusa multiplex f. silverstripe was chosen for this study, highlighting its ecological importance and unique position in leaf color diversity research.

We restructured the Materials and Methods section for better coherence, including detailed information on plant materials and growth conditions to aid replication (L. 62-77).

In the Results section, we rearranged segments for logical flow and added a table detailing key gene expression changes, increasing result transparency (L. 211).

The Discussion was streamlined to focus more on key findings, with redundant descriptions about chlorophyll's role removed, and two new tables were introduced to support discussions on enzyme roles in ALA biosynthesis and sugar metabolism pathways (L. 295, 357).

We condensed the Conclusion to succinctly summarize main findings and briefly propose future research directions, addressing the issue of length (L. 362-372).

We hope these revisions meet your expectations and improve the manuscript's quality. Thank you again for your constructive suggestions, and we look forward to any further feedback.

Round 2

Reviewer 4 Report

Comments and Suggestions for Authors

Accept in present form