Regulation of Maize Kernel Carbohydrate Metabolism by Abscisic Acid Applied at the Grain-Filling Stage at Low Soil Water Potential
Round 1
Reviewer 1 Report
Presented manuscript refers to studies concerning the effect of ABA application on carbohydrate metabolism during drought stress in maize kernels at grain-filling state. The results are interesting however basic. They are another good example for regulatory role of ABA on multiple plant physiological processes during drought stress.
The introduction is consistent, briefly describes the research problem. Although some more information about genes which encoded Suc translocated enzymes might be completed, especially that their expression were measured and presented.
Materials and Methods section should be strongly improved:
- L105-106: previous experiments should be referenced or described
- Method for measurement of glucose and fructose content should be added.
- L129: plant tissue used for analysis should be compatible within all manuscript. Also preparation of tissue used for analysis should be described more carefully
- The method of determining of sucrose invertases activities should be described in more detail, especially the procedure for preparing specific fractions for analysis.
- L153 – what is NI abbreviation mean?
Legend for Fig6 should be changed. Tissue type used for gene expression analysis is not the same as mention in the text of manuscript.
In Discussion section L335-337: the authors compare the activities of SS and INVs despite of both enzymes activities are expressed in different units. In my opinion, it is not appropriate in this case, until units are standardized.
More information is also needed about INVs encoded genes isoforms which have been analyzed. The discussion of that point of research is confused and should be improved.
Author Response
Thank you very much for your advices，I've taken them and worked very hard to correct errors.
1.Previous experiments have been described.
2. Method for measurement of glucose and fructose content have been added.
3. Preparation of tissue used for analysis have been described carefully.
4. The method of determining of sucrose invertases activities have been described in detail.
5.NI has been replaced by CIN.
6.Legend for Fig6 has been changed.
7. The comparison of SS and INVs activity has been corrected .
8.Discussion has been corrected.
Please see the attachment. Thank you again .
Author Response File: 
Author Response.pdf
Reviewer 2 Report
Presented manuscript “Regulation of Maize Kernel Carbohydrate Metabolism by Abscisic Acid Applied at the Grain-filling stage at Low Soil Water Potential” is dealing with drought stress and abscisic acid which relation well known. Drought stress dramatically change whole metabolism inside plants and abscisic acid plays important role in the regulation of metabolism. Authors particularly observed sugar metabolism of maize during grain-filling stage under drought stress. Sucrose content, sucrose synthetase activity, sucrose invertase activity were analyzed together with expression analysis of related genes. Yield of grains was also evaluated. Article novelty is not high in meaning of drought stress or ABA effects, but brings new information about sucrose metabolism in such conditions.
Particular comments
L23-24 please re-formulate sentence “yield index at 10 days” may confuse readers, it should be something like “with the lowest yield index at 10th day of stress.“
Hypothesis should be stated more clearly at the end of introduction
Authors stated that they measured Sucrose invertase but they did not provide details for cell wall invertases, vacuolar invertase, and cytoplasmic invertase measurments. This is needed to be extra explained.
In gene expression measurement, housekeeping gene/genes are no specified. Also I am not sure whether MIQE guidelines were followed. In current for it looks like simple comparison of Cq to the Cq of control on non-stressed plants, but ΔΔCq require first comparison of each Cq to housekeeping gene Cq to obtain ΔCq and then comparison of these ΔCqs to ΔCq of control on non-stressed plants.
Fisher LSD test requires ANOVA before use, and can be used only if ANOVA was significant, thus it need to be checked and corrected if needed.
Results are stylized into easy to read barcharts, which seems to be fine
In figure 6, the last sub-figure has wrong labels on X axis
In figure 6, what is the Y axis, fold changes or 2fold changes?
Article must be significantly improved mainly in material and method description.
Author Response
Thank you very much for your advices. I have fully adopted them and worked very hard to correct errors.
1."Yield index at 10 days" has been corrected.
2.Hypothesis have been stated at the end of introduction.
3. Details for cell wall invertases, vacuolar invertase, and cytoplasmic invertase measurments have been added.
4.The results of gene expression were supported by biotechnology company, and the whole process was in accordance with MiQE guidelines.
5.ANOVA has been added.
6.In figure 6, the wrong labels on X axis have been corrected.
7.In figure 6, the Y axis is 2fold changes.
Please see the attachment. Thank you again.
Author Response File: Author Response.pdf
Reviewer 3 Report
This manuscript was about study the effect of ABA on sucrose-metabolizing enzyme activities under drought stress.
- I suggest the authors to replace the Figure 1 with a higher resolution version figure.
- I am curious about the effect of ABA on the cellulose synthase activity which use udp-glucose to synthesize cellulose. Could the authors add such data?
Author Response
Thank you very much for your suggestion. I fully accept it and try my best to correct errors.
1.Figure 1 has been replaced.
2. Due to the epidemic situation, I can't go back to school for the experiment. I'm very sorry that I can't complete this modification. But in the future research, I will pay attention to its effect on plant physiology.
Please see the attachment. Once again, thank you very sincerely.
Author Response File: Author Response.pdf
Round 2
Reviewer 1 Report
The Authors took into account the reviewer’s comments. They improved ‘Material and Methods’, added missing information. However there is still one minor point that need correction. In L144 the plant tissue mention as taking for analysis should be changed to the correct tissue. The Authors should read the manuscript carefully to ensure that there are no other such errors.
With the above exception the manuscript is suitable for publication.
Author Response
Thank you very much for your advices. I adopted them and worked very hard to revise them.
In L144 the plant tissue mention as taking for analysis have been changed to the correct tissue.
Thank you very much again.Please see the attachment.
Author Response File: Author Response.pdf
Reviewer 2 Report
In previous round of review I asked for details of gene expression analysis, however method description were not changed. From the answer of authors I understand that analysis was done by external company/institution. Please state it clearly in manuscript.
It is still absolutely needed to improve description of methods. Primarily, house-keeping gene need to be stated. Correction of qPCR efficiency need to be stated if used. You can obtain this information from external institution that performed analysis.
I am not sure whether authors know what I am requesting, but without this information, article cannot be published.
for more info
https://www.gene-quantification.de/pfaffl-rel-quan-book-ch3.pdf
http://miqe.gene-quantification.info/
Other changes made in manuscript seems to fulfill requirements.
Author Response
Thank you very much for your advices.I accept them all and try my best to correct them.
1.The company has been stated.
2.House-keeping gene has been stated.
3. Correction of qPCR efficiency was not used because the comparative analysis was based on relative quantification rather than absolute quantification.
Thank you very much again!Please see the attachment.
Author Response File: Author Response.pdf
Round 3
Reviewer 2 Report
Now, it is fine
