A Mutation of cyOsPPDKB Affects Starch Structure and Gel Consistency in Rice
Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsThe manuscript “A Novel Mutation of OsPPDKB Affect Starch Structure and Gel Consistency in Rice” deals with the study of starch structure and gel consistency in rice where a Mutation was made in OsPPDKB gene. The manuscript is well-written and synthesised. However, there is scope for improvement of the manuscript before the final decision is made.
Comment
· The text mentions that rice quality is important, but it doesn't provide any background information on why rice quality matters in the context of food security or consumer preferences.
· There is a need for a more detailed explanation of the methodology employed to obtain the flo4-6 allele variant. What were the specific mutagenesis techniques used?
· The text mentions "minor genes controlling CEQ," but it doesn't explain what these genes are or how they influence rice quality.
· There is no information on the timeframe of the study, including how long it took to obtain the flo4-6 allele variant and conduct the subsequent analyses. Kinldy provide it.
· The passage mentions a significant difference in various parameters (e.g., starch granules, gel consistency) between the flo4-6 variant and the wild type. However, it does not provide details on the statistical methods used for these comparisons. It's essential to include statistical analyses to support the significance of the observed differences.
· There is no information about the statistical methods used to analyze the data, such as ANOVA or regression analysis. This should be mentioned in the Statistical Analysis subsection under the material and method heading.
· Ln 208-212: Please rewrite the line with bettter clarity.
· The significance of the 14th exon of FLO4-6 in relation to rice quality should be explained in more detail.
· The manuscript should be proofread for grammatical and typographical errors to improve clarity and readability.
· There is no conclusion section in the manuscript. Kindly provide the conclusion.
Comments on the Quality of English Language
Minor editing of English language required
Author Response
For research article
A Novel Mutation of OsPPDKB Affect Starch Structure and Gel Consistency in Rice
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Response to Reviewer 1 Comments
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1. Summary |
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Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions in the re-submitted files |
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2. Questions for General Evaluation |
Reviewer’s Evaluation |
Response and Revisions |
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Does the introduction provide sufficient background and include all relevant references? |
Yes |
Thank you for your affirmation |
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Are all the cited references relevant to the research? |
Yes |
Thank you for your affirmation |
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Is the research design appropriate? |
Yes |
Thank you for your affirmation |
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Are the methods adequately described? |
Can be improved |
We have improved the Material and Method parts |
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Are the results clearly presented? |
Can be improved |
We have improved the Results parts |
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Are the conclusions supported by the results? |
Yes |
Thank you for your affirmation |
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3. Point-by-point response to Comments and Suggestions for Authors |
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The manuscript “A Novel Mutation of OsPPDKB Affect Starch Structure and Gel Consistency in Rice” deals with the study of starch structure and gel consistency in rice where a Mutation was made in OsPPDKB gene. The manuscript is well-written and synthesised. However, there is scope for improvement of the manuscript before the final decision is made.
Comments:
Comments 1: The text mentions that rice quality is important, but it doesn't provide any background information on why rice quality matters in the context of food security or consumer preferences.
Response 1: Thank you for your reminder. We have added a description in the introduction that people's demand for rice quality is becoming increasingly strict after the increase in rice production. See lines 31-36 for details in newly submitted paper.
Comments 2: There is a need for a more detailed explanation of the methodology employed to obtain the flo4-6 allele variant. What were the specific mutagenesis techniques used?
Response 2: We have added a full description of the EMS mutagenesis and mutant screening process in the Materials and Methods section. See lines 127-133 for details in newly submitted paper.
Comments 3: The text mentions "minor genes controlling CEQ," but it doesn't explain what these genes are or how they influence rice quality.
Response 3: Wx is the major gene affecting AC and GC, and SSIIa is the major gene controlling GT. These two genes are also currently used by breeders to improve rice CEQ, as described in lines 59-71. We have added the main sites of these two genes controlling corresponding traits in the text, as shown in lines 64 and 71.
Comments 4: There is no information on the timeframe of the study, including how long it took to obtain the flo4-6 allele variant and conduct the subsequent analyses. Kinldy provide it.
Response 4: Thank you for your advice. We have added the process of QJ101 mutagenesis and flo4-7 acquisition in line 127-132 of materials and methods, and added the process of F2 acquisition for fine mapping in line 204-205.
Comments 5: The passage mentions a significant difference in various parameters (e.g., starch granules, gel consistency) between the flo4-6 variant and the wild type. However, it does not provide details on the statistical methods used for these comparisons. It's essential to include statistical analyses to support the significance of the observed differences.
Response 5: Thank you for your suggestion. We have indicated the statistical method in the figure legend of Figure 2. Please refer to line 254 of the newly submitted paper for details.
Comments 6: There is no information about the statistical methods used to analyze the data, such as ANOVA or regression analysis. This should be mentioned in the Statistical Analysis subsection under the material and method heading.
Response 1: Thank you for your advice. We have added statistical methods to the haplotype analysis section of the Materials and Methods, please refer to line 216-218 of the newly submitted paper for details, and also indicated the corresponding comparative analysis methods in the figure legends of Figures 1, 2, and 6.
Comments 7: Ln 208-212: Please rewrite the line with bettter clarity.
Response 7: Thank you for your suggestion. We have changed the description of the corresponding location. Please refer to line 229-233 of the newly submitted paper for details.
Comments 8: The significance of the 14th exon of FLO4-6 in relation to rice quality should be explained in more detail.
Response 8: We have added the description of the CDS with A as 1 at ATG and the mutation at position 2575. Thank you very much for your advice, which helped us discover an error in the first draft. The actual mutation site is at physical position 19718940. The error in the corresponding location has been corrected, and the newly uploaded Figure 6 has also been corrected.
Comments 9: The manuscript should be proofread for grammatical and typographical errors to improve clarity and readability.
Response 9: Thank you for your advice. We have made multiple grammar and error corrections throughout the text. Please refer to the parts marked in yellow for details.
Comments 10: There is no conclusion section in the manuscript. Kindly provide the conclusion.
Response 10: Thank you for your suggestion. We have added a conclusion section. Please refer to line 417-427 of the newly submitted paper for details.
Author Response File: 
Author Response.pdf
Reviewer 2 Report
Comments and Suggestions for AuthorsPlease find the attachment.
Comments for author File: Comments.pdf
No significant issues were detected. Some minor edits can improve the clarity of the English language.
Author Response
For research article
A Novel Mutation of OsPPDKB Affect Starch Structure and Gel Consistency in Rice
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Response to Reviewer 2 Comments
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1. Summary |
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Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions in the re-submitted files |
||
|
2. Questions for General Evaluation |
Reviewer’s Evaluation |
Response and Revisions |
|
Does the introduction provide sufficient background and include all relevant references? |
Yes |
Thank you for your affirmation |
|
Are all the cited references relevant to the research? |
|
Thank you for your affirmation |
|
Is the research design appropriate? |
Yes |
Thank you for your affirmation |
|
Are the methods adequately described? |
Can be improved |
We have improved the Material and Method parts |
|
Are the results clearly presented? |
Yes |
Thank you for your affirmation |
|
Are the conclusions supported by the results? |
Yes |
Thank you for your affirmation |
|
3. Point-by-point response to Comments and Suggestions for Authors |
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A Novel Mutation of OsPPDKB Affect Starch Structure and Gel Consistency in Rice
Comment 1: Must include sampling method, number of replicates, etc., for each experiment (starch content determination, amylose content determination, etc.) separately.
Response 1: We have added the acquisition method and repetition times of the samples in the figure legends of Figure 1, 2. See lines 238, 253 for details in newly submitted paper.
Comment2: Line 64-76: Use equation to show calculations.
Response 2: Thank you for your advice. We have shown the various indicators of RVA in equations. See lines 74-78 for details in newly submitted paper.
Comment3: Line 111-122: This paragraph is more likely an abstract. Please replace the current paragraph with a new paragraph that outlines your objectives instead of focusing on results.
Response 3: Thank you for your reminder. We have replaced the paragraph to highlight the purpose and innovation, and revised and added this paragraph to the conclusion section. See lines 120-123 and 417-427 for details in newly submitted paper.
Comment 4: Line 120, 125: Is there a specific reason for italicizing 'indica' and 'japonica'? If the words are not scientific names, then they should not be italicized.
Response 4: Thank you for your correction. We have removed the italics from those two words and corrected similar errors in the rest of the text.
Comment 5: Line 126: Briefly explain the grain quality of grandparents and parents (measured parameters in this study) used to create F2 population/s.
Response 5: Thank you for your reminder. We have expressed the source of the F2 population in the " Molecular cloning of FLO4-7" section of the materials and methods section. See lines 204-205 for details in newly submitted paper.
Comment 6: Line 128: Add the latitudes and longitudes of the location.
Response 6: Thank you for your reminder. We have added the longitude and latitude of the location. See lines 135-136 for details in newly submitted paper.
Comment 7: Line 131-133: Please include a reference.
Response 7: We have cited a reference here.
Comment 8: Line 131-167: Please consider adding subheadings to distinguish between the different experiments in the document. It can be challenging to follow and comprehend without them.
Response 8: Thank you for your reminder. We have added subheadings for each experiment to make the entire experimental method easier to understand.
Comment 9: Line 169: Can you clarify your process for selecting samples for microscopic examination?
Response 9: Thank you for your reminder. We have added the process of sample selection. See lines 187-202 for details in newly submitted paper.
Comment 10: Line 173: Explain 'DAF'.
Response 10: Thank you for your reminder. We have added the extension of “DAF”. See line 192 for details in newly submitted paper.
Comment 11: Line 223: Add figure number
Response 11: Thank you for your reminder. We have added the number 2 in the corresponding position.
Comment 12: Line 223-224: Move the sentence 'Previous studies … mutant' to the discussion.
Response 12: We have moved this sentence to the discussion section. See line 398 for details in newly submitted paper.
Comment 13: Line 307: Explain how you used Turkey's t-test to evaluate five haplotypes.
Response 13: First, arrange all the GC averages in descending order. The minimum average GC is Hap_3, marked as “a”. Compare this average with other averages, and label those with insignificant differences as “a”, including Hap_1, 2, and 5, until the average with significant differences, Hap_4, is labeled as “b”. Again, using the average of Hap_4 as the standard, and comparing it with each average that is smaller than it, all those that are not significant are labeled as “b”, including Hap_1, 2, and 5. The comparison of this study was completed in this way, and only the average GC between Hap3 and Hap4 showed a significant difference at the P=0.05 level.
Author Response File: Author Response.pdf
Reviewer 3 Report
Comments and Suggestions for AuthorsThis manuscript, agronomy 2633548, studied on a mutation of OsPPDKB for its effects on endosperm appearance, physical properties of flour, and also its SNP analysis was conducted. I think this study in itself was sound and its results may provide a useful information for the breeding of cooking and eating quality. I cannot detect any serious problems as a scientific article in this experiment.
However, the mutant allele at cyOsPDDKB in this study has been already detected in He et al., Agronomy 13, 2023, 1306 (doi.org/10.3390/agronomy1305136). This article of He et al. (2023) obviously has a priority for the detection of this allele. Therefore, the mutant allele in this study was not a “novel”, but a “already detected one”. I concluded that this manuscript should be thoroughly revised, particularly for its title, Abstract, Introduction, Results and Discussion, referring to the precedence study of He et al. (2023) carefully and adequately, and reconsidered again. I believe, on the other hand, that the present manuscript certainly includes a novel information, particularly for the effects of this mutant allele on gel consistency and other properties, and also SNP analysis, which were not studied in He et al (2023). Therefore, Materials and methods sections may not be affected seriously.
If the authors will withdraw this manuscript from Agronomy and submit to other journal, the same serious problems as above should be resolved.
The minor problems to be corrected are as follows:
L58: What is GT? Gelatinization temperature? This was not found in Line 48. And why ASV was not explained here?
L72: ECQ => CEQ (?)
L102: (PPDK, EC 2.7.9.1)
L112: FLO4-6 is a wild type allele of cyOsPPDKB. I think “Flo4” for the wild type allele is better.
L112: cyOsPPDKB (Os05g0405000, LOC_Os05g33570)
L173: 7 days after flowering
L182: “flo4-6” should not be italic in this line.
L196, 231: “t” should be italic.
L203: “flo4-6” should not be italic in this line.
L205, 207, 220: compared with
L210: Figure 1g-i
L218: “flo4-6” should not be italic in this line.
L223: Figure 2d, e
L231: “P” should be italic.
L232: “flo4-6” should not be italic in this line.
L263: “flo4-6” and “cyOsPPDKB” should not be italic in this line.
L268: chromosome 5,
L271: 95 kb
L277: cyOsPPDKB
L290: “cyOsPPDKB” should not be italic in this line.
L294: I think this SNP position may be indicated by the position from the top of the CDS (“A” in “ATG” is “0”).
L307: “P”
L309: “flo4-6” should not be italic in this line.
L311: Delete a space between “White-“ and “back”.
L321: Is the “FLO4-4” identical to the “FLO4-6”? If so, they should be expressed as “wild type allele (or Flo4) at cyOsPPDKB”. And also, “mutation of cyOsPPDKB” should be replaced from “mutation of FLO4-6”, which are found in many parts of this manuscript.
L324: AGPS2b
L356: AC => amylose “C” in “AC” means “content”.
L367: Delete “content”.
L380,381,383: FLO4-6 => cyOsPPDKB (?)
Author Response
For research article
A Novel Mutation of OsPPDKB Affect Starch Structure and Gel Consistency in Rice
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Response to Reviewer 3 Comments
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1. Summary |
|
|
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Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions in the re-submitted files |
||
|
2. Questions for General Evaluation |
Reviewer’s Evaluation |
Response and Revisions |
|
Does the introduction provide sufficient background and include all relevant references? |
Must be improved |
We have improved the Introduction part |
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Are all the cited references relevant to the research? |
Must be improved |
We have added the paper of flo4-6. |
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Is the research design appropriate? |
Yes |
Thank you for your affirmation |
|
Are the methods adequately described? |
Yes |
Thank you for your affirmation |
|
Are the results clearly presented? |
Can be improved |
We have improved the Results part |
|
Are the conclusions supported by the results? |
Can be improved |
We have added the Conclusion part |
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3. Point-by-point response to Comments and Suggestions for Authors |
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This manuscript, agronomy 2633548, studied on a mutation of OsPPDKB for its effects on endosperm appearance, physical properties of flour, and also its SNP analysis was conducted. I think this study in itself was sound and its results may provide a useful information for the breeding of cooking and eating quality. I cannot detect any serious problems as a scientific article in this experiment.
Comment 1: However, the mutant allele at cyOsPDDKB in this study has been already detected in He et al., Agronomy 13, 2023, 1306 (doi.org/10.3390/agronomy1305136). This article of He et al. (2023) obviously has a priority for the detection of this allele. Therefore, the mutant allele in this study was not a “novel”, but a “already detected one”. I concluded that this manuscript should be thoroughly revised, particularly for its title, Abstract, Introduction, Results and Discussion, referring to the precedence study of He et al. (2023) carefully and adequately, and reconsidered again. I believe, on the other hand, that the present manuscript certainly includes a novel information, particularly for the effects of this mutant allele on gel consistency and other properties, and also SNP analysis, which were not studied in He et al (2023). Therefore, Materials and methods sections may not be affected seriously.
If the authors will withdraw this manuscript from Agronomy and submit to other journal, the same serious problems as above should be resolved.
Response 1: It is our oversight in not noticing this article published in Agronomy in 2023. The flo4-6 in this study is different from the reported flo4-6 mutation location, and is a different allelic mutation. We have changed “flo4-6” in this paper to “flo4-7” and referenced the reference” Characterization of flo4-6, a Novel cyOsPPDKB Allele Conferring Floury Endosperm Characteristics Suitable for Dry-Milled Rice Flour Production”.
Comment 2: The minor problems to be corrected are as follows:
L58: What is GT? Gelatinization temperature? This was not found in Line 48. And why ASV was not explained here?
Response 2: Thank you for the correction. We have replaced ASV with GT in line 51 and added a description of ASV in line 66.
Comment 3: L72: ECQ => CEQ (?)
Response 3: Thank you for the correction. We have changed ECQ to CEQ.
Comment 4: L102: (PPDK, EC 2.7.9.1)
Response 4: Thank you for your revision. We have made the corresponding changes to line 109 of the newly submitted paper.
Comment 5: L112: FLO4-6 is a wild type allele of cyOsPPDKB. I think “Flo4” for the wild type allele is better.
Response 5: Thank you for your reminder. We have changed the relevant description. Please refer to lines 117-119 of the newly submitted paper for details.
Comment 6: L112: cyOsPPDKB (Os05g0405000, LOC_Os05g33570)
Response 6: Thank you for your revision. We have added the gene number in line 117.
Comment 7: L173: 7 days after flowering
Response 7: Thank you for your reminder. We have added the extension of “DAF”.
Comment 8: L182: “flo4-6” should not be italic in this line.
Response 8: Thank you for your correction. We have corrected similar errors on each title.
Comment 9: L196, 231: “t” should be italic.
Response 9: Thank you for your correction. We have made modifications in the corresponding position
Comment 10: L203: “flo4-6” should not be italic in this line.
Response 10: Thank you for your suggestion. We have corrected similar errors on each title.
Comment 11: L205, 207, 220: compared with
Response 11: Thank you for your correction. We have made the changes in the corresponding locations.
Comment 12: L210: Figure 1g-i
Response 12: Thank you for your correction. We have made the changes in the corresponding locations.
Comment 13: L218: “flo4-6” should not be italic in this line.
Response 13: Thank you for your suggestion. We have corrected similar errors on each title.
Comment 14: L223: Figure 2d, e
Response 14: Thank you for your correction. We have made the changes in the corresponding locations.
Comment 15: L231: “P” should be italic.
Response 15: Thank you for your correction. We have made the changes in the corresponding locations.
Comment 16: L232: “flo4-6” should not be italic in this line.
Response 16: Thank you for your suggestion. We have corrected similar errors on each title.
Comment 17: L263: “flo4-6” and “cyOsPPDKB” should not be italic in this line.
Response 17: Thank you for your suggestion. We have corrected similar errors on each title.
Comment 18: L268: chromosome 5
Response 18: Thank you for your correction. We have made the changes in the corresponding locations.
Comment 19: L271: 95 kb
Response 19: Thank you for your correction. We have made the changes in the corresponding locations.
Comment 20: L277: cyOsPPDKB
Response 20: Thank you for your correction. We have made the changes in the corresponding locations.
Comment 21: L290: “cyOsPPDKB” should not be italic in this line.
Response 21: Thank you for your suggestion. We have corrected similar errors on each title.
Comment 22: L294: I think this SNP position may be indicated by the position from the top of the CDS (“A” in “ATG” is “0”).
Response 22: We have added the description of the CDS with A as 1 at ATG and the mutation at position 2575. Thank you very much for your advice, which helped us discover an error in the first draft. The actual mutation site is at physical position 19718940. The error in the corresponding location has been corrected, and the newly uploaded image 6 has also been corrected.
Comment 23: L307: “P”
Response 23: Thank you for your correction. We have made the changes in the corresponding locations.
Comment 24: L309: “flo4-6” should not be italic in this line.
Response 24: Thank you for your suggestion. We have corrected similar errors on each title.
Comment 25: L311: Delete a space between “White-“ and “back”.
Response 25: Thank you for your correction. We have made the changes in the corresponding locations.
Comment 26: L321: Is the “FLO4-4” identical to the “FLO4-6”? If so, they should be expressed as “wild type allele (or Flo4) at cyOsPPDKB”. And also, “mutation of cyOsPPDKB” should be replaced from “mutation of FLO4-6”, which are found in many parts of this manuscript.
Response 26: Thank you for your advice. We have made corrections in the corresponding position in this text.
Comment 27: L324: AGPS2b
Response 27: Thank you for your advice. We have made corrections in the corresponding position.
Comment 28: L356: AC => amylose “C” in “AC” means “content”.
Response 28: Thank you for your correction. We have changed the “AC” here to "amylose”.
Comment 29: L367: Delete “content”.
Response 29: Thank you for your advice. We have made corrections in the corresponding position.
Comment 30: L380,381,383: FLO4-6 => cyOsPPDKB (?)
Response 30: Thank you for your advice. We have made corrections in the corresponding position.
Round 2
Reviewer 1 Report
Comments and Suggestions for AuthorsThe authors made significant changes in the manuscript.
Author Response
Thank you
Reviewer 3 Report
Comments and Suggestions for AuthorsThe revised manuscript v2 designated the mutation of cyOsPPDKB in this study as “flo4-7”, instead of the previous “flo4-6”. Certainly, these two mutants were derived from different original cultivars and different mutagenesis. However, they were identical for their CDS and mutated bases (and also altered amino acids). Therefore, I strongly believe that the mutation of cyOsPPDKB in this study is substantially “flo4-6”. Readers also believe the same conclusion if the revised manuscript will be published as the present form. Therefore, the authors should be revised again as my comments of the first version of manuscript.
If on the other hand, the editor of Agronomy permits the authors’ designation (“flo4-7”) because of the different origin from flo4-6 in He et al. (2023), I cannot help withdrawing my opinion for the above problem. (Probably it will be.) However even in this case, I strongly request the authors to revise at least the following matters:
1 Is “flo4-7” the mutation and/or mutant name (flo4-7 mutant line or individual), or sequence variant and/or mutant allele name (flo4-7 allele)? For example, this is mutant allele in Line 121, but in Line 140, Line 193, Line 200, Line 213, etc., this is mutant line or individual. The authors should revise not to confuse readers.
2 Delete “novel” in the title (Line 2).
3 Insert a sentence, “This mutant allele flo4-7 was identical to the flo4-6 [31] for their mutated bases (G to A) and their positions, although they derived from different original cultivars and different mutagenesis.”, for example, after Line 306 and also Line 419 (after “…endosperm.” ) to show definitely that flo4-7 and flo4-6 showed identical sequences.
Other minor problems remained are as follows:
Line 117, Line 309 and Figure 5 (a): According to the gene nomenclature system for rice (McCourch, Rice 1 (2008): 72-84), locus name (locus ID) should be non-italic (Os05g0405000, LOC_Os05g33570).
Line 121: flo4 => cyOsPPDKB
Line 139, 164, 177, 187, 191, 198: These lines should be italic.
Line 273: FLO4-7 => flo4-7
Line 275: FLO4-7 => cyOsPPDKB locus or wild type allele at cyOsPPDKB
Line 363: FLO4-7 => cyOsPPDKB
Line 395: Delete “flo4-7”.
Line 419: Delete “FLO4-7 is an allele gene of cyOsPPDKB.”
Figure 6 (a): This figure represents reverse direction compared with Figure 5 (a) (3’ to 5’). So that this is very confusing for readers. Fix the Figure 6 (a) (figure and table) as Figure 5 (a). Also in the table of Figure 6 (a), the base positions should be from “A” in the start codon. Why did the Haplotype 5 show “Y (T or C)” at position 19718753? If T, this is Hap 4, and if C, this is Hap 1.
Author Response
Please see the attachment
Author Response File: Author Response.pdf
