Genome-Wide Identification, Characterization, and Expression Analysis of the Geranylgeranyl Pyrophosphate Synthase (GGPPS) Gene Family Reveals Its Importance in Chloroplasts of Brassica oleracea L.
, Zhiyuan Fang 2, Na Zhang 2, Limei Yang 2, Yangyong Zhang 2, Mu Zhuang 2, Honghao Lv 2, Jialei Ji 2 and Yong Wang 2,*
Round 1
Reviewer 1 Report
Dear Editor,
I hope this message finds you well. I have completed the evaluation of the manuscript titled "Genome-wide identification, characterization and expression 2 analysis of the geranylgeranyl pyrophosphate synthase（3 GGPPS）gene family reveals its importance in chloroplasts of 4 Brassica oleracea L." submitted to Agriculture. After a thorough review, I would like to provide my final report on the manuscript.
Strengths:
The manuscript presents a thorough analysis of the GGPPS gene family in B. oleracea, which is relevant and helpful to the field of plant biology and genetics. The authors have successfully recognized ten GGPPS genes and investigated their phylogenetic relationships, chromosomal distribution, gene structure, and cis-acting elements. The qRT-PCR expression analysis provides a valuable understanding of the differential expression of GGPPS genes in different tissues, particularly in relation to the mutant Boas1.
Areas for Improvement (Minor Revisions Required):
1- The manuscript would benefit from a more in-depth interpretation of the results. For example, discussing the potential functional roles of the identified GGPPS genes based on their expression patterns and subcellular localization.
2- Might the authors address the discrepancy between the qRT-PCR expression results and the RNA-seq dataset analysis and provide a clear explanation for the observed differences?
3-Adding a discussion on the biological implications of the findings would enhance the significance of the study and its relevance to the broader field.
4- It is suggested to discuss the limitations of the study and potential avenues for future research in the Conclusion section.
Overall, the manuscript is well-presented, and the conclusions are of interest to the readership. The minor revisions indicated above would further improve the clarity and impact of the study.
I advise that the authors be given the chance to revise the manuscript based on the feedback provided. Once the modifications are made, I would be delighted to re-evaluate the manuscript to ensure that all the proposed modifications have been adequately addressed.
Thank you for considering my review of the manuscript. Please do not hesitate to reach out if you have any further inquiries.
Sincerely,
Author Response
Response to Reviewer Comments
Point 1: The manuscript would benefit from a more in-depth interpretation of the results. For example, discussing the potential functional roles of the identified GGPPS genes based on their expression patterns and subcellular localization.
Response 1: Thank you very much for your suggestion. I have added the corresponding explanation in the discussion section of the manuscript.
Point 2: Might the authors address the discrepancy between the qRT-PCR expression results and the RNA-seq dataset analysis and provide a clear explanation for the observed differences?
Response 2: This question is very meaningful. Before conducting the experiment, our expectation was that the results of the analysis of both the qRT-PCR and RNA-seq datasets would show consistency. Since the transcript level changes according to the time of development, the timing of RNA extraction can affect results. Both siliques and leaves were sampled 4 weeks after pollination, and the seeds in the siliques were removed at the same time. As for why the samples were taken 4 weeks after pollination, it was because the siliques of the mutants matured after 4 weeks, and the seeds also matured, and the traits of albino siliques were stable. However, the timing of sampling when performing RNA-seq experiments may not agree with mine, and it is also uncertain whether the seeds in the siliques were removed during the sampling process. The materials for RNA-seq experiments are also different from ours. This potential difference in sampling procedures could be the reason for the inconsistency between qRT-PCR expression results and analysis of RNA-seq datasets.
Point 3: Adding a discussion on the biological implications of the findings would enhance the significance of the study and its relevance to the broader field.
Response 3: Thank you very much for your suggestion. I have added the corresponding explanation in the discussion section of the manuscript.
Point 4: It is suggested to discuss the limitations of the study and potential avenues for future research in the Conclusion section.
Response 4:
Thank you very much for your suggestion. In this study, we performed bioinformatics, qRT-PCR, and subcellular localization analysis on the BoGGPPS, to investigate the role of BoGGPPS in cabbage. While no transgenic experiments were performed on the ggpps gene to determine whether there was a corresponding edited phenotype, we present a viable avenue for future research, some genes can be verified as target genes for later transgenics experiments. As a result, certain genes identified in this study hold the potential to be validated as target genes through subsequent transgenic experiments. These future experiments have the potential to shed light on the functional significance of BoGGPPS and its impact on cabbage development and physiological processes.
Thank you very much for your comments. The specific revisions have been incorporated into the new version of the manuscript, and you can check it in the new manuscript. your feedback has been instrumental in improving the quality of our research!
Sincerely,
Longxiang Yan
Reviewer 2 Report
In this paper, the authors presented a well-structured paper that explores the importance of GGPPS genes in cabbage. They successfully identified the gene family, analyzed phylogenetic relationships, studied CREs, and assessed protein structure. Additionally, they investigated gene expression using both RNA-seq and qRT-PCR, while confirming cellular localization through GFP. I believe this paper merits publication after addressing minor revisions.
1. Genes IDs can be renamed to BoGGPPS.
2. Line 58, the authors stated in their objectives that they identified albino mutant lines, this can be misleading and instead mention that the study have “used” the albino lines and mentioned the source of the mutant line.
3. In the phylogenetic tree, genes can be divided into subfamilies or sub-groups
4. The authors can also analyze the domain structure of the gene.
5. Line 335, please state the subgrouping in the result section also.
Overall, this work contributes valuable insights into the GGPPS gene family in cabbage and after addressing the above-mentioned observations, this paper would be a valuable addition to the journal.
Dear Editor,
I had the privilege of reviewing the article titled "Genome-wide identification, characterization and expression analysis of the geranylgeranyl pyrophosphate synthase（GGPPS）gene family reveals its importance in chloroplasts of Brassica oleracea L.”. In this paper, the authors presented a well-structured paper that explores the importance of GGPPS genes in cabbage. They successfully identified the gene family, analyzed phylogenetic relationships, studied CREs, and assessed protein structure. Additionally, they investigated gene expression using both RNA-seq and qRT-PCR, while confirming cellular localization through GFP. I believe this paper merits publication after addressing minor revisions.
1. Genes IDs can be renamed to BoGGPPS.
2. Line 58, the authors stated in their objectives that they identified albino mutant lines, this can be misleading and instead mention that the study have “used” the albino lines and mentioned the source of the mutant line.
3. In the phylogenetic tree, genes can be divided into subfamilies or sub-groups
4. The authors can also analyze the domain structure of the gene.
5. Line 335, please state the subgrouping in the result section also.
Overall, this work contributes valuable insights into the GGPPS gene family in cabbage and after addressing the above-mentioned observations, this paper would be a valuable addition to the journal.
Author Response
Response to Reviewer Comments
Point 1: Genes IDs can be renamed to BoGGPPS.
Response 1: Thank you very much for your suggestion. I have renamed the gene ID for GGPPS in cabbage to BOGGPPS.
Point 2: Line 58, the authors stated in their objectives that they identified albino mutant lines, this can be misleading and instead mention that the study have “used” the albino lines and mentioned the source of the mutant line.
Response 2: Thank you very much for your comments. I have added an explanation of the albino lines to the manuscript.
Point 3: In the phylogenetic tree, genes can be divided into subfamilies or sub-groups
Response 3: Thank you very much for your suggestion, I have redrawn the phylogenetic tree and divided it into three sub-groups.
Point 4: The authors can also analyze the domain structure of the gene.
Response 4: Thank you for your valuable suggestion. I have made the updates to Figure 5 in the manuscript by adding the domains section, and I have also included an analysis of the domains in the results.
Point 5: Line 335, please state the subgrouping in the result section also.
Response 5: Thank you for your input. I have included a description of the sub-groups in that paragraph.
Thank you very much for your comments. The specific revisions have been incorporated into the new version of the manuscript, and you can check it in the new manuscript. your feedback has been instrumental in improving the quality of our research!
Sincerely,
Longxiang Yan
Reviewer 3 Report
The study of GGPPS genes in cabbages reported in this study is interesting. However, the transcriptomic study was not well consistent. The growth condition and the time of RNA isolation are 2 basic factors that affect the RNA levels should be explained. The author did not explain them. Also the comparison between transcripts founds in this work with RNA seq in database should be justified and well interpreted. I fact, the difference of conditions and times of sample collection affect the RNA levels in organs. The English of the paper should be improved since many grammar structures are difficult to read. Here some points that should be addressed before publication.
Line 57: Please add one phrase about the interest of Cabbage for human.
Line 58-64: Please rephrase the sentence in order to present well the scientific context and the objective of the study.
Line 117: the growth conditions of the four cabbages used for RNA extraction were not presented. Please describe briefly the growth condition adopted. Since the transcript level changes according to the time of development, the time point of RNA extraction should be mentioned.
Line 168: the distribution and syntenic relationships of GGPPS genes in fig 3 are difficult to read. Please increase the fig size and resolution.
Line 227: this expression “To gain insights into » is used many times. Please avoid this kind of general words in scientific contexts.
Line 234: please replace “expression levels » with « transcripts level » because not all RNA will be translated to proteins.
Line 261-204: the same idea was presented in tow different phrases. Please and correct.
Line 279: replace “Relative expression” with “Transcript level” in Fig 9.
Line 356-359: the first sentence is repetition of M&M without deducing conclusions in realtio to phylogenetic relationship, chromosomal position, collinear relation, …. Please be more specific and avoid general expression..
The quality of Figures 5, 6, and 9 should be improved (size and resolution).
the english should be revised and improved
Author Response
Response to Reviewer Comments
Point 1: Line 57: Please add one phrase about the interest of Cabbage for human.
Response 1: Thank you for your suggestion. I added a description of the interest of Cabbage for human to the manuscript.
Point 2: Line 58-64: Please rephrase the sentence in order to present well the scientific context and.
Response 2: Thank you for your suggestion. I have rephrased the sentences with specific background and the objective of the study.
Point 3: Line 117: the growth conditions of the four cabbages used for RNA extraction were not presented. Please describe briefly the growth condition adopted. Since the transcript level changes according to the time of development, the time point of RNA extraction should be mentioned.
Response 3: Thank you for your suggestion. I have added the growth conditions and RNA extraction time points for the four cabbage materials into the manuscript.
Point 4: Line 168: the distribution and syntenic relationships of GGPPS genes in fig 3 are difficult to read. Please increase the fig size and resolution.
Response 4: Thank you for the suggestion. I have updated Figure 3 with a clearer specification.
Point 5: Line 227: this expression “To gain insights into » is used many times. Please avoid this kind of general words in scientific contexts.
Response 5: Thank you for the suggestion. I have revised the repetitive duplicate phrasing in the manuscript.
Point 6: Line 234: please replace “expression levels » with « transcripts level » because not all RNA will be translated to proteins.
Response 6: Thank you for the suggestion, I have changed "expression level" to "transcription level" in the manuscript as appropriate.
Point 8: Line 279: replace “Relative expression” with “Transcript level” in Fig 9.
Response 8: Thank you for your suggestion. I have replaced “Relative expression” with “Transcript level” in Figure 9.
Point 7: Line 261-204: the same idea was presented in two different phrases. Please and correct.
Response 7: Thank you for your suggestion. I have corrected these two sentences.
Point 9: Line 356-359: the first sentence is repetition of M&M without deducing conclusions in realtio to phylogenetic relationship, chromosomal position, collinear relation, …. Please be more specific and avoid general expression.
Response 9: Thank you for your suggestion. Thanks for your advice. I have added more details to the conclusion.
Point 10: The quality of Figures 5, 6, and 9 should be improved (size and resolution).
Response 10: The size and resolution of Figures 5, 6, and 9 has been improved.
Thank you very much for your comments. The specific revisions have been incorporated into the new version of the manuscript, and you can check it in the new manuscript. your feedback has been instrumental in improving the quality of our research!
Sincerely,
Longxiang Yan
Round 2
Reviewer 3 Report
The manuscript has been sufficiently improved according to the reviewers comments.
Dear Editor,
The manuscript has been sufficiently improved according to the reviewers comments.
