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All remaining issues indicated by the reviewer were addressed and revised manuscript is acceptable now.
[# PeerJ Staff Note - this decision was reviewed and approved by Paula Soares, a PeerJ Section Editor covering this Section #]
Plese address remaining issues indicated by the reviewer and make the necessary corrections in the manuscript.
1. Line 27: Replace the following line “There were six SjCCoAOMT members and fifty-two SjCOMT members in the genome of S. japonica, which was unevenly distributed on 11 chromosomes” with “There are six SjCCoAOMT members and fifty-two SjCOMT members in the genome of S. japonica, which are unevenly distributed on 11 chromosomes”
2. Line 31: “Since cepharanthine are mainly synthesized in root, SjCOMT11, SjCOMT13, SjCOMT15, SjCOMT21, SjCOMT33, SjCOMT37, and SjCOMT45 which were significantly expressed in the expression profile were further validated using qRT-PCR”. This line should be rewritten as “Significantly enhanced expression of SjCOMT11, SjCOMT13, SjCOMT15, SjCOMT21, SjCOMT33, SjCOMT37, and SjCOMT45 suggested these OMTs are essential for cepharanthine synthesis in the S. japonica roots.”
3. Line 34: “The high expression of these SjCOMTs in root suggested that they might be potential candidates involved in the synthesis of cepharanthine in S. japonica.” This line should be removed.
4. Line 83: “The main objective of this research was to analyze the characterization and function of the OMT genes family,…” should be replaced with “The main objective of this research was to analyze and characterize the function of the OMT genes family,…”
5. Line 104: “After that, all putative proteins were sent to NCBI..” should be rewritten as “ Further, all putative proteins were sent to NCBI..”
6. Line 107: “Proteins which did not match well in blast-p were send to Softberry (http://linux1.softberry.com/) for correct and proteins without target domain were manually removed.” should be rewritten as Proteins with non-significant similarity were sent to to Softberry (http://linux1.softberry.com/) for correction and proteins without target domain were manually removed.”
7. Line 146 : “All samples were performed in three biological replicates.” This line should be rewritten as “Three biological replicates per sample were used for the analysis.”
8. Line 153: “The log2 (FPKM+1) values were used to normalized and the expression heat map was generated by TBtools.” should be rewritten as “The log2 (FPKM+1) values were used for normalization and the expression heat map was generated by TBtools.”
9. Line 199: “Based on the sequence similarity and topology structures of the phylogenetic tree, OMT proteins were classed into two major subfamilies, including CCoAOMT and COMT”. Could authors explain what does this line mean?
10. Line 216: “while several SjCOMT proteins only contained four or five motifs such as SjCOMT2, SjCOMT4, SjCOMT5, SjCOMT19, SjCOMT21, SjCOMT25, SjCOMT37, and SjCOMT46.” Should be rewritten as “while SjCOMT proteins including SjCOMT2, SjCOMT4, SjCOMT5, SjCOMT19, SjCOMT21, SjCOMT25, SjCOMT37, and SjCOMT46 have four or five motifs”
Authors should be careful to not to italicize the protein names as in the above text.
11. Line 255: “RAN polymerase” should be replaced with “RNA polymerase”
12. Line 266: “This finding suggested that the OMT genes in S. japonica might be controlled by light.” should be rewritten, as light is known to regulate almost most of the genes in the plants. Authors can rewrite it as “These findings suggest these identified motifs might be important for light mediated regulation in S. japonica”.
1. Line 30: “The expression profile revealed differential gene expression patterns in five tissues”. It is not clear which OMT members are authors stating about. Line should be rewritten by mentioning which OMTs are differentially expressed in which tissues.
2. Line 88: “The stems, leaves, roots, and shoots of S. japonica”. Authors should state the age of the plants here.
3. Line 232: “We examined the expression patterns of SjCCoAOMTs and SjCOMTs in the root, bud, flower, leaf, and stem based on RNA-seq data”. Which RNA Seq data are authors mentioning about? Since it is from RNA Seq analysis deposited in NCBI, it should be referenced here with GenBank accession number.
4. Line 249: “SjCOMT11 and SjCOMT13 were barely expressed in detected tissues” What is meant by this line?
5. Line 289: “In this work, sequence similarity searches of several plant genomes enabled the identification of 58 OMTs that were unevenly distributed on the chromosomes in S. japonica”. Since, in this research work, OMT genes were identified from the already NCBI deposited data while protein sequences were retrieved on the basis of Arabidopsis OMT proteins, why authors then mentioned about several plant genomes.
No comments
No comments
Please address the concerns of the reviewers and amend the manuscript accordingly.
**PeerJ Staff Note:** Please ensure that all review, editorial, and staff comments are addressed in a response letter and any edits or clarifications mentioned in the letter are also inserted into the revised manuscript where appropriate.
1. Only the scientific names of organisms and gene names should be italic throughout the manuscript, but not the gene family names or proteins.
2. Use the complete Scientific names once in the manuscript, then use abbreviations like A. thaliana for Arabidopsis thaliana.
3. In abstract, add a bit more information about the qRT-PCR, which genes were selected for this and why.
4. Expand the introduction to provide sufficient background information on Stephania japonica, cepharanthine, and O-methyltransferase gene family. Clearly state the research gap and the significance of the study in the context of existing literature.
5. Ensure that the methods section is well-structured and detailed enough for readers to replicate the study. Describe the data sources, experimental procedures, and analytical tools with clarity.
6. Present the results in a clear and organized manner. Use tables and figures to illustrate key findings and trends. Ensure that all relevant data is included and properly analyzed.
7. In discussion, provide a comprehensive interpretation of the results, relating them to the research objectives. Discuss the implications of the findings and their relevance to cepharanthine biosynthesis or potential applications.
8. Ensure that all sources are properly cited, and the reference list is accurate and consistent. Follow the journal's citation style guidelines.
9. Keep the quality of the images to or above 300dpi and add a bit more details about the figure in the figure legends.
1. Line 78: "sequences of the S. japonica could be acquired" should be "sequences of S. japonica were acquired."
2. Line 81: "utilized" should be "used."
3. Line 87: "regarded as the OMT gene family members in S. japonica and renamed for further analysis according to their relative positions on chromosomes" is confusing. Please rephrase this sentence.
4. Line 99: Why use the Neighbor-joining (NJ) method instead of Maximum-likelihood or the UPGMA?
5. Line 119/120: Provide references for the “comparative Ct (22-··Ct) method” and “GAPDH as the internal reference”.
6. Line 128: The selection of the promoter sequence up to 1400 bp is noteworthy. However, it is common in many studies to consider a longer promoter region, usually around 2000 bp. Can the authors provide a rationale for choosing this specific length of 1400 bp for the promoter analysis in this study? Justification for this choice will strengthen the significance of the findings related to the cis-acting elements.
Results:
1. Line 162: The word "Amon" is likely a typo, it should be "Among."
2. Line 166-174: Information about the Orthologs and paralogs should be added to the text.
3. Only the genes names and scientific names should be in italics.
Discussion:
1. Line 250: "anti-coronavirus activity" should be defined or referenced.
2. Line 255: "58 OMTs that were unevenly distributed on the chromosomes in S. japonica, which might be related to proximal gene duplication" - Please specify the evidence supporting the claim of proximal gene duplication.
3. Line 261: "CCoAOMT and COMT were the two major subfamilies of OMTs" - Please specify the full names of "CCoAOMT" and "COMT" at first use.
4. Line 266: "Exploring the expression profiles of SjCCoAOMT and SjCOMT in five tissues is necessary to provide insight" - Please specify the reason for exploring expression profiles in this context. "SjCCoAOMT and SjCOMT in five tissues is necessary" - Add a comma after "SjCCoAOMT" for grammatical correctness.
Conclusions:
1. Line 285: "The comprehensive study of phylogenetic relationship" should be "Comprehensive study of --"
Figures:
ANOVA analysis should be applied to quantitative data, e.g. Figure 5
Overall, the manuscript provides valuable insights into the O-methyltransferase gene family in Stephania japonica. However, there are some areas that need clarification and minor improvements. The authors should address the highlighted issues and consider the comments for enhancing the clarity and scientific rigor of the paper.
No comment
1. Authors have analyzed the expression of different OMTs in various tissues but have not mentioned the age of the plant when the tissue was sampled. It should be added in the legend for Figure 5 and also in material & methods section.
Liang et al discuss the identification and expression analysis of OMT protein family in Stephania japonica. Authors have performed gene structure, phylogenomic analysis, motif analysis, structure analysis, and gene expression analysis to speculate the functions of OMT gene family. The manuscript has explored the characteristics of OMT genes through sequence alignments and other bioinformatics tools.
1. SjCOMT28, 34, 47, 5, and 9 have no transcript presence in any collected tissue while SjCOMT24 is only present in root. It is interesting to note that mere presence of gene doesn’t signify its role in overall development. It also put a question mark to incorporate the selected protein in phylogenetic analysis.
2. The expression analysis is not aligned with the transcriptome data like the expression of representing the SjCOMT15 where the expression of target gene is higher in stem instead of leaf. Another one is SjCOMT45. The selection criteria to shortlist SjCOMTs for expression analysis is also not clear.
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