Transcriptome Insights into Carbohydrate Metabolism and Frying Quality Traits in Waxy and Mealy Potatoes

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Transcriptome Insights into Carbohydrate Metabolism and Frying Quality Traits in Waxy and Mealy potatoes has been reported. We are grateful for valuable suggestions and comments on our manuscript. I agree that these are exciting research topics. However, the manuscript must be crucible-revised before it meets precise research requirements. Here, I express the main reasons for Major Revision and suggest how the authors can improve them.

1.  Lines 147-149 on page 4, "doubled monoploid potato genome DM 1-3516 R44" was selected as the reference gene for downstream analysis of DEGs. However, reference [20] did not indicate whether the potatoe S. tuberosum Group Phureja was a mealy or a waxy potato. Since this gene is not highly conserved, please explain the rationality of using this gene. It is suggested that the same type of potatoes be used for omics analysis and comparison to find a common reference genome, which would be more appropriate.

2. Line 139 on page 4, In Table 1, Two “Sample name” was labeled "Chuback" under Group A. Is this a typographical error, or was this duplication intentional in the experimental design? Please verify and clarify.

3. Lines 170-189 on page 5, In Section 2.5, 726 carbohydrate metabolism-related genes were selected based on KEGG, paralogs from Solanum/Nicotiana, and annotations from the reference genome (DM 1–3 516 R44). However, in Section 3.5, 168 genes showed significant differences between the two potato types. The authors should explicitly state whether these 168 genes are annotated in the DM 1–3 516 R44 genome. If these genes are absent or poorly represented in the reference genome, the validity of using this genome for comparative analysis would be questionable. As important analytical parameters, please be sure to explain reference genome uniformly and clearly.

4. Lines 263-281 on page 7, In Section 3.4 Functional classification of DEGs, the GO enrichment analysis revealed no significant terms related to starch metabolism in the upregulated DEGs of either potato type. Did the authors specifically compare DEGs between mealy and waxy potatoes for starch metabolism-related pathways? If no significant terms were identified, does this imply that starch metabolism is not a key differentiating factor between the two types, despite their distinct starch compositions (higher amylose in mealy vs. amylopectin in waxy)? Please discuss this discrepancy.

5. Line 282 on page 8, Figure 3. The text is too small and blurry. It is suggested to improve the clarity.

Author Response

Dear Reviewer,

We sincerely appreciate your thoughtful and detailed feedback. Your comments have been instrumental in improving the clarity and scientific rigor of our manuscript.

Below, we provide a section-by-section summary of the major revisions made in response to your suggestions.

 

Comment: Lines 147-149 on page 4, "doubled monoploid potato genome DM 1-3516 R44" was selected as the reference gene for downstream analysis of DEGs. However, reference [20] did not indicate whether the potatoe S. tuberosum Group Phureja was a mealy or a waxy potato. Since this gene is not highly conserved, please explain the rationality of using this gene. It is suggested that the same type of potatoes be used for omics analysis and comparison to find a common reference genome, which would be more appropriate.

Response 1: We agree that the reference genome does not have a clearly defined as mealy or waxy. However, we selected this reference genome for below reasons.

The DM 1-3 516 R44 genome is a doubled monoploid, which allows for high-quality, chromosome-scale assembly without the complications of heterozygosity. This enables more accurate alignment and gene annotation in transcriptome analysis compared to other reference genomes.

Also, this genome has been widely adopted as the reference in most potato genomic studies to date. Its public availability make it a practical and reproducible choice for comprehensive interpretation of results.

 

Comment: Line 139 on page 4, In Table 1, Two “Sample name” was labeled "Chuback" under Group A. Is this a typographical error, or was this duplication intentional in the experimental design? Please verify and clarify.

Response 2: We have corrected the typographical error and clarified that one of the entries labeled as “Chubaek” should be “S18051-5”. We apologize for the oversight and appreciate your attention to detail. [page 4. 149-150.]

 

Comment: Lines 170-189 on page 5, In Section 2.5, 726 carbohydrate metabolism-related genes were selected based on KEGG, paralogs from Solanum/Nicotiana, and annotations from the reference genome (DM 1–3 516 R44). However, in Section 3.5, 168 genes showed significant differences between the two potato types. The authors should explicitly state whether these 168 genes are annotated in the DM 1–3 516 R44 genome. If these genes are absent or poorly represented in the reference genome, the validity of using this genome for comparative analysis would be questionable. As important analytical parameters, please be sure to explain reference genome uniformly and clearly.

Response 3: We confirm that all carbohydrate-metabolism related genes were verified through BLAST and are present in the reference genome. The full list of genes and the subset of expressed genes are provided in Table S1 and Table S3, respectively.

To improve clarity, we have also added a statement in the manuscript indicating that these gene lists are available in the supplementary data. [page 5. 195.][page 10. 364-365.]

 

Comment: Lines 263-281 on page 7, In Section 3.4 Functional classification of DEGs, the GO enrichment analysis revealed no significant terms related to starch metabolism in the upregulated DEGs of either potato type. Did the authors specifically compare DEGs between mealy and waxy potatoes for starch metabolism-related pathways? If no significant terms were identified, does this imply that starch metabolism is not a key differentiating factor between the two types, despite their distinct starch compositions (higher amylose in mealy vs. amylopectin in waxy)? Please discuss this discrepancy.

Response 4: As the reviewer correctly noted, starch metabolism is a key trait distinguishing mealy and waxy potatoes. The absence of significant GO terms related to starch metabolism is likely due to the different genetic background among the potato genotypes used. So to compensate these, we conducted mapping on each starch-related gene. We specifically examined the expression levels of 81 genes related to starch metabolism, independent of the GO enrichment results. These genes were analyzed and visualized in revised Figure 6A, and their expression patterns regarding phenotypes were discussed in detail in the Discussion section. [page 12. 379-385.][page 18. 489-607.]

To clarify this point, we have added an explanation in the Discussion section, highlighting the potential impact of genetic background on GO enrichment outcomes and justifying our decision to analyze starch metabolism genes individually. [page 22. 674-692.]

 

Comment: Line 282 on page 8, Figure 3. The text is too small and blurry. It is suggested to improve the clarity.

Response 5: To improve the figure clarity, we limited GO term visualization in Figure 4 to the top 10 terms per category, while the full GO terms are available in Table S4. [page 9. 308-313.]

 

We once again thank the reviewer for the valuable feedback and suggestions, which have helped us substantially improve the quality and clarity of our manuscript.

Reviewer 2 Report

Comments and Suggestions for Authors

Dear Authors,

Congratulations on this excellent and comprehensive research! Your work presents a thorough transcriptomic investigation into the molecular basis of frying quality in potatoes, particularly distinguishing between waxy and mealy cultivars. The integration of gene expression, and metabolic pathway analysis is commendable and adds substantial value to both academic research and practical applications in potato breeding.

I will give you some general comments and suggestions for improvement:

• The abstract is informative but quite dense. Consider slightly reducing complexity or breaking long sentences to improve readability for a broader audience.

• The manuscript is well organized; however, some transitions between sections (especially from Results to Discussion) could be smoother with short summary sentences or bridge paragraphs.

• The discussion could be enhanced by more explicitly linking gene expression patterns with their physiological consequences during frying. For instance, connecting specific genes to texture formation or color more clearly.

• Statistical Robustness: In sections with small sample sizes (e.g., one replicate for group B), consider discussing the limitations this imposes on statistical inference.

• References: You reference many studies well, but the addition of a brief comparison with recent omics-based potato breeding studies could provide stronger positioning of your contribution.

Specific suggestions:

• Line 22–24: When introducing coexpression with bZIP transcription factors, mention how this connects to known regulatory mechanisms (e.g., ABA signaling) earlier for better context.

• Figure 8: It's a strong summary figure. Adding brief captions for each subpanel (A–D) could improve immediate understanding.

• Methods Section: The RNA-seq processing and DEG criteria are rigorous. It may be helpful to briefly justify the log2FC ≥ 1.0 threshold biologically, not just statistically.

Author Response

Dear Reviewer,

We sincerely appreciate your thoughtful and detailed feedback. Your comments have been instrumental in improving the clarity and scientific rigor of our manuscript.

Below, we provide a section-by-section summary of the major revisions made in response to your suggestions.

 

Comment: The abstract is informative but quite dense. Consider slightly reducing complexity or breaking long sentences to improve readability for a broader audience.

Response 1: Thank you for your suggestion. We have revised the abstract by breaking down long sentences and simplifying complex structures to enhance clarity and accessibility for a broader audience, while retaining the core scientific content. [page 1. 12-29.]

 

Comment: The manuscript is well organized; however, some transitions between sections (especially from Results to Discussion) could be smoother with short summary sentences or bridge paragraphs.

Response 2: To improve the flow, we have added a brief bridge paragraph at the start of the Discussion section, addressing the needs to interpret the results more thoroughly in the Discussion section. [page 18. 483-488.]

 

Comment: The discussion could be enhanced by more explicitly linking gene expression patterns with their physiological consequences during frying. For instance, connecting specific genes to texture formation or color more clearly.

Response 3: We fully agree with this point. The revised Discussion now includes how does the specific gene expression network (SS1, SS3, SuSy4, invertases, bZIP transcription factors, and SnRK2.4) associate with their physiological consequences during frying, specifically color differences through Maillard reaction. [page 20. 587-594.]

 

Comment: Statistical Robustness: In sections with small sample sizes (e.g., one replicate for group B), consider discussing the limitations this imposes on statistical inference.

Response 4: We have added a statement in Abstract, Discussion, and Conclusion section discussing the limitation of biological replication in group B and small size of group A with different genetic background, and how it may affect the statistical inference, particularly for differential gene expression and GO enrichment analyses. We also emphasized the need for future studies with expanded biological replication. [page 1. 12-29.] [page 22. 674-692.] [page 23. 696-707.]

 

Comment: References: You reference many studies well, but the addition of a brief comparison with recent omics-based potato breeding studies could provide stronger positioning of your contribution.

Response 5: We highlight our positions as a unique transcriptome research in frying-related studies at the Introduction section. This statement emphasizes the gap in transcriptomic studies comparing mealy and waxy types, and justifies the rationale for our investigation. [page 3. 97-115.]

 

Comment: Line 22–24: When introducing coexpression with bZIP transcription factors, mention how this connects to known regulatory mechanisms (e.g., ABA signaling) earlier for better context.

Response 6: This has been revised. The association between bZIP transcription factors and ABA signaling pathways is now mentioned to provide better biological context and improve reader comprehension. [page 1. 22-25.]

 

Comment: Figure 8: It's a strong summary figure. Adding brief captions for each subpanel (A–D) could improve immediate understanding.

Response 7: We have relabeled the Figure 8 as Figure 6, and added brief informative captions to each subpanel to enhance clarity and facilitate quick interpretation. [page 12. 379-385.]

 

Comment: Methods Section: The RNA-seq processing and DEG criteria are rigorous. It may be helpful to briefly justify the log2FC ≥ 1.0 threshold biologically, not just statistically.

Response 8: We have added a brief justification in the Methods section, noting that log2FC ≥ 1.0 was chosen to reflect biologically meaningful changes in gene expression. [page 4. 164-167.]

 

We once again thank the reviewer for the valuable feedback and suggestions, which have helped us substantially improve the quality and clarity of our manuscript.

Reviewer 3 Report

Comments and Suggestions for Authors

See the file

Comments for author File: Comments.pdf

Comments on the Quality of English Language

See the file

Author Response

Dear Reviewer,

We sincerely appreciate your thoughtful and detailed feedback. Your comments have been instrumental in improving the clarity and scientific rigor of our manuscript.

Below, we provide a section-by-section summary of the major revisions made in response to your suggestions.

 

Abstract

Comment: add more details for the abstract. This study conducted transcriptome analysis on two types of potatoes with distinct end-use purposes to identify gene expressions related to desirable frying qualities, focusing on texture and appearance after frying. Add more details.

Comment: This study provides a foundational understanding for breeding potato cultivars with improved processing quality and identifies key targets for marker-assisted breeding to enhance frying-related quality. Add limits and new avenues for researchers.

Response: We have revised the abstract by including more specific details about the identified genes and their potential mechanisms and functions. To improve readability, we separated long sentences into shorter, clearer ones. Additionally, we included a brief mention of the study’s limitations and proposed directions for future research. [page 1. 12-29.]

 

Introduction

Comment: Potato (Solanum tuberosum L.), a member of the Solanum genus, ranks among the world’s four major food crops, sustaining over a billion people worldwide. Originating from South America, potatoes are now extensively cultivated because of their agricultural efficiency and high yield potential. Potatoes develop underground storage organs known as tubers, which facilitate asexual propagation through tiny buds called eyes, enabling efficient and straightforward cultivation. Moreover, potatoes demonstrate a high yield per unit area relative to other major crops and have a short growing cycle of approximately 100 days, making it possible to harvest multiple planting cycles within a year. Add references.

Comment: Mealy potato cultivars, including “Russet Burbank,” “Shepody,” and “Atlantic,” are primarily used to produce French fries and potato chips, which are highly valuable in the processed food industry. Potato chips, made with thinly sliced potatoes, hold a dominant position in the global snack market, with a value exceeding $35 billion. References.

Response: We added appropriate references to support the previously unreferenced statements, thereby strengthening the introductory statements [1-3][4,5]. [page 1. 38-42.][page 2. 56-57.]

 

Comment: improve the introduction. Clarify the problematic, support the ideas with references and examples.

Response: The introduction has been revised to better articulate the research problem. It now provides a clearer explanation of the importance of frying-related traits in potatoes, highlights the gap in transcriptomic studies comparing mealy and waxy types, and justifies the rationale for our investigation. Relevant references and examples have been incorporated throughout to support key arguments and provide context. [page 3. 97-115.]

 

Material and Methods

Comment: Plant materials and frying quality-related phenotype assessment. Add figure or table to summarize the materials (in addition to table 1).

Comment: please add statistical analysis.

Response: We revised the Table 1 by separating information related to plant materials from phenotypic results, thereby improving clarity and structure. [page 4. 149.]. In response to the reviewer’s comment, we also added a detailed description of the statistical methods used for phenotypic data analysis, including the comparison of frying traits between different potato types. [page 3. 134-135.]

 

Results

Comment: Phenotypes related to frying quality. Add table or figure.

Comment: The four A-group A potato samples chosen for the transcriptome analysis displayed distinct traits related to frying qualities. Did compare the results.

Comment: The two mealy potatoes exhibited a lower glucose 222 concentration, with “Dubaek” and “EarlyFry showing” displaying 0.10 and low (below the detection limit), while the two waxy potatoes exhibited higher concentrations of 0.40 224 and 0.50. compare statistically.

Response: We added the frying quality traits of B-group potatoes and presented the data in a new Table 2. [page 6. 242.]. Statistical analyses comparing mealy and waxy potatoes across A and B-group samples were added, and the results are visualized using boxplots in the revised Figure 1. [page 7. 245-249.]

 

Comment: (Table 1): please avoid to combine results and materials in the same table.

Response: Plant materials and results are now clearly separated. [page 6. 242.]

 

Comment: Figure 1. Unclear (correlation matrix).

Response: Figure 2B (correlation matrix) has been updated with higher resolution for clarity. [page 7. 262-268.]

 

Comment: Figure 3. G. unclear legend.

Response: To improve the figure clarity, we limited GO term visualization in Figure 4 to the top 10 terms per category, while the full GO terms are available in Table S4. [page 9. 308-313.]

 

Comment: Figure 8: displace it to results.

Response: The previous Figure 8 has been repositioned to the Results section and renumbered as Figure 6. [page 12. 379-385.]

 

Discussion

Comment: please combine results and discussion in the same section.

Response: Thank you for your suggestion. While we fully understand the rationale behind integrating the Results and Discussion, we chose to retain them as separate section in this manuscript. This decision was made because the Discussion includes in-depth interpretation and functional insights that go beyond the scope of the direct results, including gene expression regarding hormonal signaling. Metabolic implications, and molecular reaction during frying processes. We believe this separation allows for a more structured and focused narrative, especially given the complexity of the transcriptome data. However, we realized necessity for a smooth transition between two sections. We have added a concise bridge paragraph at the start of the Discussion. This paragraph addresses the needs to interpret the results more thoroughly in the Discussion section, thereby enhancing the logical flow without merging the two sections. [page 18. 483-488.]

 

Conclusion

Comment: mention the limits of the study and then new avenues.

Response: We added the limits of the study and then new avenues likewise other sections of this paper, considering the limited sample size, and to suggest future directions for research. [page 23. 696-707.]

 

We also revised the manuscript to correct typographical errors and improve English grammar and readability.

 

We once again thank the reviewer for the valuable feedback and suggestions, which have helped us substantially improve the quality and clarity of our manuscript.

 

Added References

1. Lutaladio, N.; Castaldi, L. Potato: The hidden treasure. Journal of food composition and analysis 2009, 22, 491-493.
2. Hijmans, R.J. Global distribution of the potato crop. American journal of potato research 2001, 78, 403-412.
3. Haverkort, A.; Struik, P. Yield levels of potato crops: recent achievements and future prospects. Field Crops Research 2015, 182, 76-85.
4. Jaggan, M.; Mu, T.; Sun, H. The effect of potato (Solanum tuberosum L.) cultivars on the sensory, nutritional, functional, and safety properties of French fries. Journal of Food Processing and Preservation 2020, 44, e14912.
5. Kirkman, M.A. Global markets for processed potato products. In Potato biology and biotechnology; Elsevier: 2007; pp. 27-44.

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript has undergone significant revisions and improvements, with more detailed content, supplemented with necessary illustrations, and a greater focus on the theme. I agree that these are exciting research topics. The manuscript is based on impressive empirical evidence and makes an original contribution. Only Minor Revisions are needed before it can be published.

1. To more clearly explain the differences between "Waxy and Mealy”, it is suggested to supplement the genetic background introduction of four types of potato samples.

2. When explaining in text and listing in tables, please unify the order of the two varieties potatoes in the text. Such as Table 2 and Table 4, the order of the two mealy samples (“Dubaek” and "EarlyFry") and the two waxy samples (“Chubaek” and "S18051-5") is kept consistent, and the order of the text descriptions is also.

3. Lines 631-637 on Page 24, It supplemented and explained the relationship between the two types of potatoes in the transmission of transcription factors, related enzymes and signal systems. This is an important conclusion and supplement, which greatly enhances the research value of this article. If the author can carry out the necessary verification work on these important genes, the credibility of the conclusion will be greatly enhanced.

Author Response

Dear Reviewer,

We sincerely appreciate your detailed feedback. Your comments have been instrumental in improving the clarity and quality of our manuscript and have provided insights for further research.

Below, we provide a section-by-section summary of the major revisions made in response to your suggestions.

 

Comment: To more clearly explain the differences between “Waxy and Mealy”, it is suggested to supplement the genetic background introduction of four types of potato samples.

Response 1: Thank you for your helpful suggestion. We agree with your comment, and to clarify the differences between the two potato types, we have added a description in Section 2.1. Plant Materials explaining how the representative mealy and waxy samples were selected. Additionally, to aid readers’ understanding regarding differences, we included an explanation of the typical characteristics distinguishing mealy and waxy potatoes, as well as measured traits of our samples, that were followed by an interpretation of the observed gene expression patterns in relation to these phenotypes. [page 3. 121-124.] [page 18. 491-500.]

 

Comment: When explaining in text and listing in tables, please unify the order of the two varieties potatoes in the text. Such as Table 2 and Table 4, the order of the two mealy samples (“Dubaek” and "EarlyFry") and the two waxy samples (“Chubaek” and "S18051-5") is kept consistent, and the order of the text descriptions is also.

Response 2: Thank you for your detailed suggestions. We have revised the manuscript to ensure that the order of the potato samples is consistent throughout the text and in Tables 1,2,3 and 4.

 

Comment: Lines 631-637 on Page 24, It supplemented and explained the relationship between the two types of potatoes in the transmission of transcription factors, related enzymes and signal systems. This is an important conclusion and supplement, which greatly enhances the research value of this article. If the author can carry out the necessary verification work on these important genes, the credibility of the conclusion will be greatly enhanced.

Response 3: Thank you for your valuable comment. We fully agree that functional validation of the transcription factors would be an important next step to further strengthen our findings. However, this work was designed as an exploratory study based on comparative transcriptomic analysis, and due to current experimental limitations, functional assays could not be conducted at this stage. Instead, we performed co-expression analysis between the transcription factors and starch synthases to explore potential functional associations, and we have supplemented the interpretation with literature-based evidence on their putative roles. We are planning follow-up studies involving yeast one-hybrid assays to investigate transcription factors and gene interactions. These future directions have been stated in the revised manuscript. [page 20. 577-580.]

 

We also revised the manuscript to correct typographical errors and improve English grammar and readability.

 

We once again thank the reviewer for the valuable suggestions, which have helped us substantially improve the quality and clarity of our manuscript.

Reviewer 3 Report

Comments and Suggestions for Authors

no comments

Author Response

Comments: no comments

Response: no comments