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Your manuscript was accepted. Congratulations
[# PeerJ Staff Note - this decision was reviewed and approved by Julin Maloof, a PeerJ Section Editor covering this Section #]
Julin Maloof, the Section Editor, has commented and said:
> I thank the authors for the revisions. There is one point that is still unclear. In the revised manuscript they state "StringTie and Ballgown (http://www.bioconductor.org/packages/release/bioc/html/ballgown.html) were used to estimate the expression levels of all transcripts and perform expression level for mRNAs by calculating fragments per kilobase of transcript per million mapped reads (FPKM) (Pertea et al. 2015). DESeq2 was used to identify DEGs from a number of important gene sets (Love et al. 2014)." This does not address my concern about input to DESeq2. Was FPKM used as input to DESeq2? If so, that is a mistake. Raw counts should be used as input to DEseq2. Please see "Input Data" in the DEseq2 manual at https://rdrr.io/bioc/DESeq2/f/vignettes/DESeq2.Rmd
Please address this in your resubmitted manuscript.
Dear authors,
Even if the reviewers offer to accept your manuscript after the changes you have made, you still need to address some editorial concerns.
"Some more info and possibly reanalysis of RNASeq data is needed
What program was used for read filtering?
The papers needs citations for "Goatools" and "GOseq" both of these have published papers to cite.
DESEQ2 requires raw read counts, not FPKM, but the methods section states that FPKM was calculated by RSEM. The analysis may need to be redone using "expected counts" output from RSEM, not FPKM.
Maize has very good annotation, far better than could be obtained by de novo assembly of the reads in this experiment. Why was de novo assembly done? Were the de novo transcripts used for read quantification or were reads mapped to a reference genome/transcriptome? And if so which one?"
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The authors have answered all my questions, and the current version of the manuscript can be accepted.
After my previous review, the authors have completed the necessary edits. Literature additions have also been made in appropriate places. Although there is a small spelling error in the references (for example, the journal name should start with a capital letter in line 496), this does not prevent the publication of this study. Figures and tables are prepared with sufficient clarity and explanation for the reader.
Research question well defined, relevant & meaningful. It is stated how research fills an identified knowledge gap.
The results and findings section is sufficiently explained. I think that this study will fill the gaps in the literature and will be a step for future studies.
Although your study was well-constructed to understand the role of transcriptional factors in salt tolerance of maize seedlings, you did not present it well. Please address the reviewers' concerns and revise your manuscript, paying attention to the points I have highlighted below.
Be sure to address each comment as you make revisions. Unfulfilled or unexplained changes may result in your manuscript being rejected.
Line 81: what length? what weight? specify root or shoot.
Line 112-114: It is not correct to give the results of the study in this section, state your purpose here.
Line 127: if you used three technical replicates, how many biological replicates were there? The reliability of studies without biological replication is questioned. Elaborate the materials and methods section in more detail.
Line 132: Are one of the cultivars you used transgenic and one wild type? You have never mentioned this before. Provide more information about the cultivars you used.
Line 196: I don't think that one-way ANOVA is the correct statistical model for this study. Please re-analyse your data according to two-way ANOVA.
Line 279: Is the number of genes you selected for qRT-PCR eight or twelve? You said twelve before.
Explain more about the qRT-PCR results and include them in the discussion. Also, the conclusion section is rather stumpy. It should be better written and key findings should be emphasised. The discussion section of the study should also be strengthened.
Note: R2 has provided an annotated PDF
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The analyses are largely insufficient, and some are irrelevant to salt stress. The authors need to perform comprehensive and in-depth analyses to explore the genes and pathways associated with salt stress.
1. The resolution of Fig.2A and Fig.5A is too low, making the contents indiscernible. Please provide high-resolution versions of these figures.
2. Please clarify which gene sets were used for the GO enrichment analysis in Fig.4. The GO enrichment analysis should be conducted separately for DEGs detected in the ST0 vs. SS0, ST3 vs. SS3, and ST7 vs. SS7 comparisons.
3. The analysis of transcription factors encoded by DEGs detected in the ST0 vs. SS0, ST3 vs. SS3, and ST7 vs. SS7 comparisons in Fig.6 is not meaningful for studying salt stress. Instead, the authors should compare the expression level changes of published transcription factors or genes known to be associated with salt stress.
The writing language can be improved. If a more academic language is used, the study will reveal itself more. The use of current literature is quite good, but the number of literature is insufficient at some points and more should be added.
Research question well defined, relevant & meaningful. It is stated how research fills an identified knowledge gap.
The methods are explained very well. But, it is not clear what is being said in some sentences. If the sentences are written in a more simplified manner, they will be understandable.
Conclusions are well stated, linked to original research question & supporting results.
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