Genome-Wide Identification of the Argonaute Protein Family and Its Expression Analysis under PEG6000, ABA and Heat Treatments in Populus alba × P. glandulosa

Round 1

Reviewer 1 Report

This study is devoted to the identification of Argonaut family proteins in poplar. The authors have done interesting work in terms of analyzing the evolution of PtAGO genes, the chromosomal localization of PtAGO, the genetic structure and analysis of regulatory elements in the promoter region of PtAGO genes. In addition, the profile of PtAGO expression in different poplar tissues and under the influence of PEG6000 and ABA was analyzed.

The authors performed a large study using the most modern research methods. This research is unique and carries great value for fundamental science. Therefore, the topic of this manuscript is relevant for Forests. However, I have some suggestions for this manuscript:

General remarks:

1)                 The authors should correct the title of the manuscript (Ms). The Ms presents the results of the expression of genes of the Argonaut family under the influence of PEG6000 (osmotic stress), ABA - stress hormone (imitation of stress in a plant), and high temperatures (heat stress). Abiotic stresses usually include: exposure to elevated and lowered temperatures, salt stress and drought. In my opinion, it is not correct to write all abiotic stresses in the title. The authors should specify under the influence of what stresses. In addition, the Latin name of the poplar should be indicated in the name.

2)                  When analyzing gene expression, the authors used only one reference gene Actin. When carrying out such results, at least two, and preferably 3 reference genes are most often used. Since under the influence of abiotic stresses, the expression of “housekeeping” genes can also change. In order for your data to be relevant, it is necessary to add expression data when using another reference gene, for example, GAPDH, UBQ or UBC.

3)                 There are no symbols of statistical validity of the data on the graphs of gene expression (letter designations). Perhaps the manuscript will look better if you present Figures 5-7 in the form of a heatmap.

4)                 The authors should add to the chapter materials and methods how different poplar fabrics were isolated, in particular how xylem and phloem were separated.

Minor remarks:

On page 1: line 14, small RNAs => sRNAs;

                   line 38, and further along the text, the space between the text and the square bracket is omitted;

                   line 39, In The leaves=> In the leaves

On page 2: line 60, add the Latin name of the poplar;

On page 3: line 103, cultured in vitro for 4 months => cultured in vitro for 4 months;

               line 123-125, add manufacturer and country;

               line 138, “Hence, these results” - check the font;

On page 4: line 146, into Group III while AGOs in M. polymorpha and => into Group III, while AGOs in M. polymorpha and;

               Table 1, add group I, II, III in the column name;

On page 7: line 187, “As for intron/extron, PtAGOs contained introns and most of them contained

more than 20 introns)...” the sentence should be rewritten;

               line 196, “involved in, cis-“ => “involved in cis-“;

Fig.5. The notation on the charts does not correspond to the notation in the description of the figure. In particular, xylem and phloem;

On page 10: line 256, The title of the chapter should be written in italics;

               line 262, “at 96 h after treat” - check the font;

References - check the font.

I recommend this Ms for publication after essential major revisions.

Comments for author File: Comments.pdf

The article is well written, small corrections are needed, which I indicated above.

Author Response

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Author Response File: Author Response.docx

Reviewer 2 Report

Dear authors,

the presented manuscript is quite valuable, and I believe that after careful editing and bringing some clarifications, it is suitable for publication.

L37 - L38 - it is necessary to edit the phrase; its meaning is not understood.

L39 - the description of the mutation should be explained more scientifically.

L60 - This statement is more suitable for the "Conclusion" section. The "Introduction" lacks a clear description of the purpose of this study, as well as the "in silico" resources used. It is not clearly explained why they resorted to studies under stress conditions (cultivation in artificial conditions induces additional stress). From what you have presented, it is clear that this manuscript describes only the first part of a much larger study; I suspect that there will also be a characterisation of the gene sequences identified in silico. I think that these steps should also be discussed/explained, so it seems a crude truncation that doesn't really make sense.

The study has value and is even an example by which already-known data can be used to increase knowledge in a certain direction.

I think some corrections to English expressions are necessary.

Author Response

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Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

The authors have significantly corrected the text according to my recommendations.

In my opinion, the authors should leave only heatmaps as figures 5-7, remove the graphs.

Perhaps Figure 8 will look better in the form of a heat map (remove graphs).

In Figure 5, the caption to the figure should be corrected. The figure shows the designations Ph and Xy, and the figure caption stands for P, phloem; X, xylem.

Author Response

Please see the attachment.

Author Response File: Author Response.docx