BrPARP1, a Poly (ADP-Ribose) Polymerase Gene, Is Involved in Root Development in Brassica rapa under Drought Stress

Round 1

Reviewer 1 Report

I just reviewed the manuscript entitled “BrPARP1, a poly (ADP-ribose) polymerase gene is involved in root development in Brassica rapa under drought stress” and to me, it seems to be a good approach.

The authors used morphological, biochemical and molecular data to justify their findings. To be honest, I was looking for some plant morphological and physiological data like growth rate, dry mass accumulation, r/s ratio that could have been more useful to support the entire findings.

However, there are some other queries that are needed to address to improve this manuscript.

Line 28: stress?

Line 77-80: Please rephrase the sentence.

Line 347: Please add the extraction procedure of the sample for proline analysis (see the example https://doi.org/10.3390/plants9111511)

Line 355: Please write the determination of NAD+ procedure in brief.

There is no information on statistical analysis…why? It should be a major concern.

Author Response

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Author Response File: Author Response.docx

Reviewer 2 Report

The work by Shi et al concerns the functional characterization of a poly(ADP-ribose) polymerase (BARP1)gene in Brassica rapa plants exposed to drought stress. Particularly, the authors demonstrated the role of BARP1 in the regulation of the plant's stress response by analyzing drought tolerance phenomena in transgenic Brassica lines obtained by targeting the BARP1 gene via RNAi technique.
Although the study provides interesting results on the potential contribution of BARP1 in the control of drought tolerance in B.rapa, more efforts should be done to improve the manuscript quality overall and to make the presented data more compelling as well as adequately supported in the Discussion section. I also warmly suggest asking help from an English proofreading service or an English native speaker, as I am confident that a carefull revision of English grammar and style would greatly improve the presentation of results. 
Finally, there are several points that need to be elucidated further, as I indicated more in detail in the following evaluation report.

l.20 INTERFERENCE

l.23 WAS

l.28 Stress

l.30 respondS

l.33-34 Some of the keywords are already present in the title of the article. I suggest changing them with alternatives and/or find new ones.

l.71 I would change part of this sentence as following proposed: PARP is one of the main energy demanding processes under stress conditions.

l.103 WAS calculated

l.103 I would change value with ratio

l.127-130 This sentence is misleading, please revise it.

l.133 The authors used PEG to induce stress here, so they should speak about osmotic rather than drought stress. The following parts should be revised accordingly.

l.144 Did the authors mean under normal osmotic conditions?

l.150-151 I got the meaning of this part, but it should be rephrased and written more clearly.

l.158 WERE also analyzed

l.162 compareD

l.163 I guess it should be IS instead of as here

l.171-172 revise the sentence from 'the the plants about...to 3 weeks'.

l.177-178 There are no letters above the bars in the graphs, did the authors mean asteriks? I suggest indicating which statistical analysis was conducted to test the significance of collected data (here as well as for other results presented in the manuscript). A paragraph dedicated to statistics should be added in the methods section.

l.200-201 Could the authors provide any empirical measurement attesting that the cell layers in the meristematic zone of RNAi plants were significantly more than in the wt?

l.203 related TO cell cycle

l.212-215 Please, provide information about statistical measurements in Figure 5B caption.

l.218-219 I suggest revising this sentence as it follows: in order to clarify the BrPARP1 silencing-mediated molecular mechanisms at the basis of the enhanced drought tolerance of transgenic B. rapa plants, ...

l.222 indicateD

l.223 this sentence is not very clear: did the authors mean that BrPARP1 could be involved in drought stress response mechanisms by interaction with stress responsive genes?

l.237 duplicationS

l.240-241 This part of the sentence has no meaning

l.249-250 This sentence is not clear.

l.251 What did the authors mean with 'WGT event could distinguish their genomes '

l.260-261 The importance of gene duplication events is redundantly expressed here, however what is not clear is why and in which terms gene duplication could be functional to future gene characterization studies.

l.268 I am quite surprised to read mannitol here...did the authors mean PEG?

l.270 lineS WERE (..unless there was one line showing a even better drought tolerance than the others)

l.273 mannitol or PEG?

l.279-282 Is there other information in the literature attesting the expression changes of these genes in B. rapa under water deficit?
My feeling is that the Discussion section should be improved by including more information/experimental evidence from the literature eventually supporting the collected results. In the actual form, the Discussion seems a replication of results description rather than a critical interpretation of the obtained data. 

l.284 Please, substitute plants with them (ie. which enables them)

l.284 I would suggest using 'improved' instead of stronger here

l.285 I would substitute the last part of the sentence with '..capacity, and thereby a higher stress tolerance behaviour'.

l. 293 ... supported our previous hypothesis..

l.293-295 This sentence is not very clear. I suggest revising the core message.

l.298-299 I suggest revising this part as it follows: ...were grown in a growth chamber under the following conditions: 16h/22°C...

l.300 relative humidity

l.315 WAS analyzed

l.328 which kind of PEG? 6000, 8000?

l.328 I would suggest specifying under OSMOTIC stress

l.329 The composition of the staining buffer should be indicated

l.334 The authors should explain the reason for which these 3 transgenic lines were chosen. Did the choice depend on the specific level of BrPARP1 downregulation in these lines? Are there any Southern blot data referred to this aspect? 

l.334 More information should be provided about the drought stress imposition. For instance: how many RNAi plants were subjected to the treatment and how many RNAi plants were used as irrigated control (I refer here to the number of biological replicates used in the experiment of course)? The same for WT plants. Was the drought stress imposed by witholding irrigation of by progressively limiting the daily amount of water provided to the plants until reaching a desired percentage? On which basis did the authors decide to impose drought for 3 weeks?

l-336-337 It is not clear how the PEG treatment was induced. I mean: was it performed on the same plants previously exposed to drought or did the authors use a different set of plants? In the second case, (as also suggested for the drought stress treatment) please, provide number of biological replicates used for each group of WT and RNAi plants subjected tot he treatment.

l.358 From which tissue was the RNA extracted? and how many mg of tissue were used for the extraction? Did the authors check RNA integrity at the Bioanalyzer, or alternatevely with other tools, before cDNA preparation?

Author Response

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Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

I thank Shi et al. for their effort in revising and updating the status of the manuscript. In the revised MS authors remarkably addressed all concerns including statistical issues. 

Author Response

Dear reviewers, dear editors,

 

Thank you very much for your comments and suggestions on this manuscript. Thank you for your hard work on this manuscript and your affirmation of this article. Please accept our sincere thanks.

Reviewer 2 Report

Dear authors,

you can find my evaluation in the attached file.

Comments for author File: Comments.pdf

Author Response

Please see the attachment.

Author Response File: Author Response.docx