Exploring the Root Morphological Traits of Diverse-Origin Cultivated Soybean

Round 1

Reviewer 1 Report

The findings of the study present interesting insightful and well-needed research, addressing a critical issue. This study is up to date, using advanced research facilities, and appears to be supported by a thorough analysis of an extensive and diverse cultivar dataset, with interesting results. I recommend this research for publication, however, I believe there are some points that need clarification and minor mistakes that should be corrected.

1.      I would suggest the author reconsider the title if it can get better, or more precise.

2.      The abstract seems a bit lengthy if the authors can make it short

3.      Line 18: remove the Brackets from Glycine max (L). Merr

4.      The author should reconsider the Abstract and write it by following a proper pattern i.e. introduction, methods, results, and the outcome or conclusion.

5.      Why did the authors select these specific traits? There are a significant number of root traits that can provide more novel insight into the research.

6.      The author should revise the introduction, as many sentences are not complete or do not make sense.

7.      Line 32: Use the complete phrase ‘plant breeders’, rather than ‘breeders’.

8.      Line 33: The word ‘enhance’ does not seem the right word to use here, please replace it with the fit word with the right context.

9.      Line 43-45: Please revise the sentence as it is uncompleted and does not make any sense.

10.   Line 48: The datasets have more of cultivars Korean origin but there seems missing the mention of the current state of soybean production or yield output of Korea in the literature review, while other countries are mentioned

11.   Line 59-62: what does the author mean to multi-stresses, as the author refers to drought stress in the next sentence? Please revise it.

12.   Line 99-105: the objective of the study is not well-defined. Please revise and clearly state the objective.

13.   Line 107: The heading “Plant Materials and Growth Condition” does not look fit. Please revise.

14.   Line 107, Section 2.1: The methodology is not clear to understand, please provide more detail.

15.   Line 109: What is the reason for using the PVC pipes for the experiment?

16.   Line 120: What type of tray was used to place the PVC pipes, and what is the reason to harvesting plants at the V2 stage?

17.   Please increase the text size of Table 3, as it's very small to read

Comments for author File: Comments.pdf

Author Response

Reviewer 1’s comments

 

I appreciate the authors for their insightful and well-needed research, addressing a critical issue. This study is up to date, using advanced research facilities, and appears to be supported by a thorough analysis of an extensive and diverse cultivar dataset, with interesting results. I recommend this research for publication, however, I believe there are some points that need clarification and minor mistakes that should be corrected.

Dear reviewer, thank you for your interest in our research and such valuable comments. We really appreciate your time and your well-needed comments on our manuscript. We have tried our level best to answer each and every comment and made changes where you have rightly suggested. All the changes are made by using the track changes feature and are highlighted with a green color. We hope that we satisfy our worthy reviewer, still if there are some changes our reviewer wants to suggest, please let us know. We thank you again, your valuable comments have made our manuscript better.

Comment 1: I would suggest the author reconsider the title if it can get better, or more precise.

Response: Dear reviewer, thank you for your suggestion. We have made changes to the title of our manuscript. As per your suggestion, we tried to make it precise while also having the core information or concept of the study.

Comments 2 and 4: The abstract seems a bit lengthy if the authors can make it short. The author should reconsider the Abstract and write it by following a proper pattern i.e. introduction, methods, results, and the outcome or conclusion.

Response: Thank you for your kind suggestion. Dear reviewer, we have made changes to our abstract per your suggestion. We have tried to make our abstract more precise and to the point. Furthermore, you have rightly suggested the missing of proper pattern in our abstract, our abstract did not had the materials and method part, and now we have added it. Now, we believe our abstract pattern is clear and in the right order. Please find the made changes from line 19-21. We hope we have met the reviewers demand in this regard. Thank you.

Comment 3: Line 18: remove the Brackets from Glycine max (L). Merr

Response: Dear reviewer, thank you for pointing out this to us. We have removed the brackets from Glycine max (L). Merr. Please track the changes in line 18. Thank you.

Comment 5: Why did the authors select these specific traits? There are a significant number of root traits that can provide more novel insight into the research.

Response: Thank you for your valuable comment. We appreciate your kind suggestion in this regard. Dear reviewer, you are on point regarding the considerable number of traits using Expression 1200XL scanner and WinRhizo software can measure. But the objective of our research was to give a precise and targeted way forward for the future research that would follow. We tried to mainly to focus on the traits that are very important and specifically have proven roles in abiotic stress and also overall growth and yield output. We did followed with a large number of traits, but after the vast amount of the reported literature on root traits and after our personal analysis, we found our current selected traits seemed to be the most important, independent and were the variables that differentiated the germplasms and, in results, their responses. Most of the other traits were seemed dependent on our selected traits, whereas principle component analysis (PCA) also proved the importance of these traits. Furthermore, a huge amount of literature is also present, whereby many repeated experiments, these traits proved to be the causing difference between the response to stresses and overall performance. So being precise and targeted, we selected these traits so the breeders can target and put efforts towards obtaining optimal root system. We hope we have satisfied our reviewer with our response. Thank you.

Comment 6: The author should revise the introduction, as many sentences are not complete or do not make sense

Response: Thank you for your valuable comment. Dear reviewer, we have made changes in our introduction part. We have tried to write the sentences with more clarity so that it can make a complete sense. Please find the changes made from line 44-50, 52-53, and in lines 71-73. Thank you.  

Comment 7. Line 32: Use the complete phrase ‘plant breeders’, rather than ‘breeders’

Response: Thank you for pointing this out. Dear reviewer, we have replaced the word with a complete phrase. Please find the changes in line 99. Thank you.

Comment 8. Line 33: The word ‘enhance’ does not seem the right word to use here, please replace it with the fit word with the right context.

Response: Thank you for your comment. Dear reviewer, we have made the suggested changes. We rewrote the paragraph and tried to put all the words in the right context. Please find the changes made in lines 34-36.

Comment 9: Line 43-45: Please revise the sentence as it is uncompleted and does not make any sense

Response: Dear reviewer, thank you for your valuable comment. We have revised the mentioned sentences to make it more clear to understand, please find it in lines 44-49 and 52-53.

Comment 10: Line 48: The datasets have more of cultivars Korean origin but there seems missing the mention of the current state of soybean production or yield output of Korea in the literature review, while other countries are mentioned.

Response: Dear reviewer, thank you so much for pointing out this point to us. Although we used germplasms originating from South Korea, including 32 others, here, we wanted to briefly give a global perspective and trend of soybean yield output. The omission of the figures of South Korea is done as we wanted to give just an overview of soybean production worldwide with just a mention of the highest producer countries (USDA, ipad.fas.usda.gov/countrysummary). If the reviewer still considers the states need to be mentioned, please let us know, we will revise it in the second round. Thank you

Comment 11: Line 59-62: what does the author mean to multi-stresses, as the author refers to drought stress in the next sentence? Please revise it

Response: Thank you for your valuable comment. Dear reviewer by ‘multistress’ we mean the occurrence of two or more abiotic stress at the same time. Abiotic stresses, specifically low or high temperature, waterlogging, drought stress high salinity, heavy metals, and ultraviolet radiations, when occur at the same time, are detrimental to crop growth and takes a huge toll on crop yield output worldwide. It is very important to equip crops with a ‘multistress’ tolerance, where in which the crop root system plays the most important role.

Comment 12: Line 99-105: the objective of the study is not well-defined. Please revise and clearly state the objective.

Response: Thank you so much for your valuable comment. Dear Reviewer, to make our objective clear, we have replaced some words with more fitting ones. Please find the objective of our study in lines 94-102. Thank you.

Comment 13: Line 107: The heading “Plant Materials and Growth Condition” does not look fit. Please revise.

Response: Dear reviewer, thank you for pointing this out to us. As per the comment, we have revised the heading. Please find the revised heading in line 104. Thank you.

Comment 14: Line 107, Section 2.1: The methodology is not clear to understand, please provide more detail.

Response: Thank you for your well-pointed comment. Dear reviewer, as per your valuable comment, we have revised our “Methodology” section. We have tried to rewrite it with more clarity and to make it more easy to reproduce or follow. Please find the changes made in lines 106, 111-121. Thank you.

Comment 15: Line 109: What is the reason for using the PVC pipes for the experiment?

Response: Thank you for your valuable comment. Dear reviewer, as we know roots have high plasticity which means they gets change or adjust with the provided medium and environment. It has been noted that experiment conducted in pots potentially limits the root length, which can result in misleading data. Sowing in pots is fine when the objectives are not root traits specific. In our case, as our core objective of the study was to comprehensively evaluate a high number of diverse germplasm datasets for the root morphological traits (RMTs), we wanted to provide such a medium where the roots express full to their potential, without any hindrance. The polyvinyl chloride (PVC) pipes used in this experiment measured 60 cm in height and 16.5 cm in diameter which provides a favorable condition for the plant roots to grow at their full potential with no hindrance. We hope we satisfy the worthy reviewer's concern. Thank you.

Comment 16: Line 120: What type of tray was used to place the PVC pipes, and what is the reason to harvesting plants at the V2 stage?

Response: Thank you for your valuable comments. Dear reviewer, your kind comment has two parts, we have answered in following a similar order below;

Part 1: The trays used in our setup are made up of thick plastic, measuring around 27 cm in height, 65 cm in length, and 30 cm in width. A total number of 40 pipes can be adjusted perfectly in one tray without disturbing the angle. The selection of this tray was done on its best carrying ability of the pipes and the potential of having finely adjustment of the pipes.

Part 2. The V2 stage, which stands for "vegetative 2," typically occurs when the plant has two fully developed trifoliate leaves. At the V2 stage, the soybean plants develop a sufficient root system to start assessing root traits effectively, thus we collected root samples on V2 stage. Another reason is that “To identify genetic diversity related to root traits during weaning period, we were assessed at the vegetative V2 growth stage, corresponding to 2–3 trifoliate leaf development.”

Comment 17: Please increase the text size of Table 3, as it's very small to read

Response: Thank you so much for your kind suggestion. Dear reviewer, the font size kept in Table 3 was because of the large size of the table. As we increase the font size by a single margin, the table splits into two pages, which makes it quite difficult to read and understand. We have tried to adjust it to a single page to make it more readable and easily understandable. Still, if our honorable reviewer thinks that the font size of the table needs to be increased, please let us know we will do it. 

Reviewer 2 Report

The manuscript titled "Exploring Root Morphological Traits in Cultivated Soybean for  Breeding Advancements: A Comprehensive Study" contains important information for science, breeding and agricultural practice. I appreciate that this is a two-year study, although a greenhouse study. However, there is little information in the text whether the research results were repeatable over the years. In greenhouse conditions, the results are usually repeatable, but this is not the case in field experiments. Many studies assess soybean varieties and compare plant biometric features, morphology, yield, seed quality, etc. The authors conducted research on the root system of various soybean varieties, including those from different regions of the world. I read the publications with interest and congratulate you on the idea. The root system determines the growth and development of crops. However, the manuscript needs improvement. I included detailed comments in the original text (pdf).

General notes:
write whether the results were repeatable over the years
add to keywords: germplasm, plant breeding
in the Introduction, write very briefly about the disadvantages of soy, e.g. anti-nutritional substances
improve especially the Material and Methods chapter
briefly describe the scale used to describe the development phases
whether fertilization, inoculation, seed treatment, and how the plants were watered were used
correct the numbering of tables and figures, including those in the appendix
why Korean varieties performed best in the tests
in the conclusion, write what further experiment are necessary
correct the literature list


I hope that my comments will allow the authors to improve the text of the manuscript. Thank you for your cooperation.

After making corrections, I recommend publishing the manuscript in the journal Agronomy

Comments for author File: Comments.pdf

Author Response

Reviewer 2’s comments:

 

The manuscript titled "Exploring Root Morphological Traits in Cultivated Soybean for Breeding Advancements: A Comprehensive Study" contains important information for science, breeding, and agricultural practice. I appreciate that this is a two-year study, although a greenhouse study. However, there is little information in the text on whether the research results were repeatable over the years. In greenhouse conditions, the results are usually repeatable, but this is not the case in field experiments. Many studies assess soybean varieties and compare plant biometric features, morphology, yield, seed quality, etc. The authors conducted research on the root system of various soybean varieties, including those from different regions of the world. I read the publications with interest and congratulate you on the idea. The root system determines the growth and development of crops. However, the manuscript needs improvement. I included detailed comments in the original text.

Response: Dear reviewer, thank you for reviewing our manuscript and recommending it for publication. Your well-pointed comments have refined our manuscript and increased its clarity and accuracy. We have tried to answer and all the comments. If further changes are needed, please let us know. Following are the responses to the worthy reviewer's valuable comments. We have performed the suggested changes with the track changes feature, also, we have highlighted the changes with the yellow color in the manuscript. Thank you.   

Comment 1: write whether the results were repeatable over the years

Response: Thank you so much for your valuable comment. Dear reviewer, we do believe that our results are reproducible, provided that the growth condition is kept the same. We performed analysis of variance (ANOVA) for both of the years, where each batch (2021-2022) of the experiment were taken as different environment, to check whether the results were consistent or reproducible. We found no significant difference between the environments i.e. 2021 and 2022, in the same germplasms in most of the important traits, i.e; total root length (TRL) (p>0.12), the average diameter (AD) (p> 0.60), root volume (RV) (p> 0.11) confirming the consistency of the results. However, other traits such as the number of tips (NT) (p>0.001), number of forks (NF) (p>0.008), and tertiary totlength (TTL) (p>0.01) were found significantly different.  

Comment 2: add to keywords: germplasm, plant breeding

Response: Thank you for the suggestion. Dear reviewer, we have added the terms ‘germplasm and plant breeding’ to the keywords. Please find it in line 38. Thank you.

Comment 3: in the Introduction, write very briefly about the disadvantages of soy, e.g. anti-nutritional substances

Response: Thank you for your valuable suggestion. Dear reviewer, we have added the part ‘Antinutrients (ANFs)’ part in the pointed section. Please find the addition in lines 47-50. Thank you.

Comment 3: improve, especially the Material and Methods chapter

Response: Thank you for pointing this out to us. Dear reviewer as per your valuable suggestion, we have completely revised the ‘ Materials and Methods’. We hope that it has become more clear and accurate and reproducible. Please find the changes made in lines 111-121. Thank you.

Comment 4: briefly describe the scale used to describe the development phases

Response: Thank you for your comment. Dear reviewer, by V2 stage, we mean when two unfolded trifoliate leaves appears [1]. As per your valuable suggestion, we have briefly added in the section what we mean by the V2 growth stage. Please find the suggested addition in line 121-122. Furthermore, the mentioned reference in your comment has defined the stages quite differently. We have followed a book, named Tennessee Soybean Production Handbook [1]. We hope it cleared our reviewers' concern, if further clarification is needed, please let us know. Thank you.

Comment 5: whether fertilization, inoculation, seed treatment, and how the plants were watered were used

Response: Dear reviewer, thank you for your valuable comment. As per the suggestion, we have revised our text and tried to add every necessary information that can be needed to reproduce our experiment. We have added the mentioned information in the section, we hope that it will help the readers. Please find the revised information in lines 118-122. Thank you.

Comment 6: correct the numbering of tables and figures, including those in the appendix

Response: Dear reviewer, we have cross-checked all the tables and figures, we believe the numbering of the tables and figures is correct. However, we made some changes which may remove the confusion regarding the numbering. Please find the changes made in lines 204, 206 and 235. However we request our reviewer, if there seems any wrong numbering, please point it out to us, we will happily correct it. Thank you.

Comment 7: why Korean varieties performed best in the tests

Response: Dear reviewer, thank you for a well-pointed comment. The reason for the difference in performance is possibly the environment of the origins of the germplasms. Our result reveals a potential role of origin in shaping RMTs and RSA. Literature has revealed high plasticity in the root system, which makes it sensitive to the environment and growth conditions. The variation of results and different trends based on origins in our experiment confirms this possibility. Furthermore, it is reported in the literature, that under water-limited conditions, the availability of water and nutrients to plants is contingent on environmental conditions, root system morphology, architectural traits, and competition; consequently, plants under such conditions tend to develop deeper root systems to access water. To further confirm this supposition, we suggest that further studies need to be conducted exploring RMTs and the potential of origins in shaping and influencing root systems.   

Comment 8: in the conclusion, write what further experiments are necessary, correct the literature list

Response: Thank you for your valuable suggestion. Dear reviewer, we have added the suggestion for the further experiments that needs to be followed. Please find the addition in lines 374-375.

 

Minor comments on the original file in pdf:

 

Comment: Were the results repeatable, where there any significant differences between the years?

Response: Thank you for your valuable comment. Dear reviewer, although our experiments are conducted in a controlled environment however the comprehensive nature of our study and the two-year setup are quite positive signs that germplasms will perform similar in field conditions too. Furthermore, no significant differences between the environment in the major traits confirm the results' reproducibility. Additionally, we would like to suggest that experiments need to be conducted to explore the potential role of the origins of the germplasm could also be explored.

Comment: ‘Table S1, Table S2, Table S3, and Figure S1’

Response: Dear reviewer, Table S1, Table S2, Table S3, and Figure S1 mentioned in lines 109, 255, 257, and 261, are provided in the supplementary materials. Please find these tables and the figure in supplementary materials. Thank you.

Comment: Correction in references; add polish.

Response: Dear reviewer, thank you for your comment. We have crosschecked this reference from different sites but found no differences. I suppose your good self was pointing out to the language that it needs to be written in the Polish language, which we found to be already in Polish. However, if you could further explain or if there is anything we can do with it please let us know, we would happily revise it.

Comment: References correction and botanical names italicize

Response: Dear reviewer, thank you for pointing out errors in the references section. We have revised the references and italicized the scientific names as well.