Integrated Transcriptomic and Metabolomic Analyses of Seed-Filling Disorders in Soybeans Under Different Ecological Conditions

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript entitled “Integrated Transcriptomic and Metabolomic Analyses of Seed-filling Disorders in Soybeans under Different Ecological Conditions” investigates the transcriptomic and metabolomic basis of seed-filling disorders in a wrinkle-prone soybean cultivar (Heihe 43) grown in a high-temperature region of China (Shenyang).

The authors report that high temperatures during the seed-filling period in Shenyang lead to the upregulation of genes involved in starch hydrolysis and sucrose metabolism, and the downregulation of genes associated with plant hormone signal transduction in Heihe 43.

While the study is interesting, I have two major concerns:

1. Since only one susceptible variety was tested, are the results sufficiently reliable?
2. As the work was conducted in only one specific region, can the findings be generalized to other environments?

In addition, given that environmental conditions strongly affect the expression of the identified genes, what practical solutions do the authors propose for addressing the problem of wrinkle-prone varieties in high-temperature regions?

We have genotype × location effect in this experiment, however there is no table result for this.

Table 1 is not the “Results” of your experiment. This type of information should be presented as Materials and Methods and in a supplementary file.

 

Other comments:

Introduction:

• Line 45: “Glycine max” in Italics.
• Lines 56-58: This paragraph is similar to the previous mentioned paragraph (Lines 49-51).
• Lines 58-60. This paragraph is also mentioned already (Lines 54-56).
• Line 74: Be sure that correctly type “sh2019 and sh2021”.
• Line 76: Names of genes should be in italics.
• Lines 126-132: These parts do not belong at the end of the 'Introduction' section. Here, you should present and discuss your hypothesis.

Materials and Methods:

• Which type of experimental design applied?

Results:

• Lines 222-226: This paragraph belongs in the Materials and Methods section.
• Line 226: “The results indicated that …”. Here you should present the results of ANOVA, not a picture!
• Section 3.2 (lines 249-257). To show a significant difference, you need statistical indicators. This paragraph belongs to the Materials and Methods not “Results”.
• Table 1: It is not “Results”. Where is “statistics”?
• Figure 2: We just need the last row of this table. The correlation of one meteorological factor with another one is not useful.

Discussion:

• Lines 476-479: This paragraph belongs to the “Results” not Discussion. You can’t refer to your results (Lines 479, 487, 488, 528, …) in Discussion section.
• Lines 497-505: Here I just see a repetition of Results not a Discussion on Results.
• Lines 509-513: Please provide reference for your sentences about peas.
• Line 534: Please provide full name of “ARF and GH3” in their first application in the text.

Conclusions:

• Lines 597-600: You need a statistical analysis (ANOVA) for this claim.

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors The manuscript investigates the potential molecular mechanisms underlying soybean seed development. Experiments conducted in Jiamusi and Shenyang revealed significant differences in sugar and starch accumulation, as well as in the activities of key enzymes such as sucrose synthase and invertase. Integrated transcriptomic and metabolomic analyses further identified candidate targets and metabolites associated with sugar metabolism and hormone pathways. The experiments are well-designed, and the findings are clearly presented. Overall, this study provides insights into the physiological and molecular basis of seed wrinkling in soybean, and significant for soybean agriculture. Below are my suggestions for further improving the manuscript.

1. The authors propose that temperature is a key factor influencing the formation of wrinkled seeds during the grain-filling stage. It would be valuable to explore whether altering temperature can partially or fully reverse the seed phenotypes. Conducting controlled greenhouse experiments accompanied by qPCR analysis of potential targets involved in sugar metabolism and hormone signaling pathways, as well as wrinkled-seed formation, could enhance the study.
2. Although temperature is initially hypothesized as a major influencing factor, the current transcriptomic and metabolomic analyses lack direct connection to temperature variation. Strengthening this logical linkage is recommended.
3. All figure legends should provide more detailed information to facilitate reader interpretation. In Figure 1A and 1B, the apparent differences in petri dish size and the number of soybean seeds shown may raise questions about whether yield differences are being compared. Please clarify these aspects.
4. For Figure 3, 7, and 8, qPCR validation of key candidate genes would strengthen the credibility of the findings by confirming their expression patterns.
5. Line 118-134 Some sentence does not appear suitable for a research paper and requires improvement. I would recommend making it more concise and employing a formal register.
6. High temperatures under climate change pose major threats to soybean yield and breeding. The authors should emphasize these challenges and the relevance of this study in the Introduction and Discussion. Future directions for developing climate-resilient soybean varieties should also be highlighted.

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The authors have made a sincere effort to address all the comments raised during the first round of review. Overall, the revision addressed all raised issues, and I do not have any further concerns. 

Author Response

Thank you for professionally and patiently putting forward suggestions that can improve the quality of this research.