Exogenous ABA Induces Osmotic Adjustment, Improves Leaf Water Relations and Water Use Efficiency, But Not Yield in Soybean under Water Stress
Round 1
Reviewer 1 Report
The manuscript is interesting and well written. However,some major changes need to be made.
Most importantly, there are very little statistical tests performed, or at least, shown. Please include more statistical details.
Figure 6 needs to be rotated.
The discussion is too long. Please condense it.
The conclusions are rather the summary of results. So what are your conclusions from the results? Are ABA applications relevant for soybean growers? What do you recommend? Please re-write the concusions.
Some minor remarks:
Check the whole manuscript carefully as there are several grammatical mistakes (for example line 62: "improve" is correct; lines 253 and 308: "withdrawn" is correct) and incorrect spellings (e.g. line 88: "experimwnt")
Table1: why did you include the abbreviations RWC, TW, PW and JD in the legend? They do not show up in the table so these explanations are not neceassary here.
Lines 244 - 245: in lines 241 you mention that the ABA was applied one day before the watering treatment, but later on you talk about three days before the watering treatment. Which is correct?
Line 380: it needs to be "Medrano, H."
Author Response
Response to Reviewer #1
1. The manuscript is interesting and well written. However, some major changes need to be made.
Authors’ response: We appreciate the positive comment of this reviewer.
2. Most importantly, there are very little statistical tests performed, or at least, shown. Please include more statistical details.
Authors’ response: We thank the reviewer for this observation and we have responded positively by conducting the statistical test, which results are included in the figures.
3. Figure 6 needs to be rotated.
Authors’ response: In the revised version we have rotated Fig. 6.
4. The discussion is too long. Please condense it.
Authors’ response: We are so grateful to the reviewer for this observation and we have re-written some parts of the discussion to condense it.
5. The conclusions are rather the summary of results. So, what are your conclusions from the results? Are ABA applications relevant for soybean growers? What do you recommend? Please re-write the conclusions.
Authors’ response: In the revised version, we have re-written the conclusions. They are now in Lines 338-342.
6. Check the whole manuscript carefully as there are several grammatical mistakes (for example line 62: "improve" is correct; lines 253 and 308: "withdrawn" is correct) and incorrect spellings (e.g. line 88: "experiment")
Authors’ response: We are so grateful to the reviewer for these observations and we have responded positively by checking the whole manuscript and correcting grammatical mistakes and typos. They are highlighted in blue in the revised version.
7. Table1: why did you include the abbreviations RWC, TW, PW and JD in the legend? They do not show up in the table so these explanations are not necessary here.
Authors’ response: In the revised version, we have deleted the abbreviations RWC, TW, PW and JD from the legend of Table 1
8. Lines 244 - 245: in lines 241 you mention that the ABA was applied one day before the watering treatment, but later on you talk about three days before the watering treatment. Which is correct?
Authors’ response: We thank the reviewer for this observation and we have corrected that inconsistency. Through the revised version it reads now that ABA was applied one day before the water treatment commenced but the pots were transfer to the growth chamber three days before water treatments (Lines 241-244).
9. Line 380: it needs to be "Medrano, H."
Authors’ response: Corrected as advised.
Author Response File: 
Author Response.docx
Reviewer 2 Report
The grammar should be improved. For instance, in line 16, please rephrase the sentence, it's better to state as 'Exogenous ABA was applied 45 days after sowing (DAS), the physiological and seed yield response of soybean to exogenous ABA were examined as the soil was drying; in line 23, the comma between 'Exogenous ABA application' and 'increased ABA' should be deleted; in line 20, 'was' after 'RWC and OA' should be changed to 'were'. Overall, please examine the grammar through the whole manuscript.
Please provide the expanded form immediately before or after the first use of abbreviation. For instance, please provide the full words for RWC and OA in the Abstract where they first time occurs.
In experiment 2, although the objective is to study the effect of exogenous application of 10 um ABA, the 0 um ABA should be also included as the control group.
Author Response
Response to Reviewer #2
Point 1: The grammar should be improved. For instance, in line 16, please rephrase the sentence, it's better to state as 'Exogenous ABA was applied 45 days after sowing (DAS), the physiological and seed yield response of soybean to exogenous ABA were examined as the soil was drying; in line 23, the comma between 'Exogenous ABA application' and 'increased ABA' should be deleted; in line 20, 'was' after 'RWC and OA' should be changed to 'were'. Overall, please examine the grammar through the whole manuscript.
Authors’ response: We thank the reviewer for these observations and in the revised version we have corrected the grammar. The revised version was checked by a native English speaker.
2. Please provide the expanded form immediately before or after the first use of abbreviation. For instance, please provide the full words for RWC and OA in the Abstract where they first time occurs.
Authors’ response: In the revised version, we had provided the expanded form of the abbreviation after the first use of it (highlight in blue).
3. In experiment 2, although the objective is to study the effect of exogenous application of 10 um ABA, the 0 um ABA should be also included as the control group.
Authors’ response: In the revised version, while describing experiment 2, we have clearly indicated that 0 um ABA was used as the control treatment.
Author Response File: Author Response.docx
Reviewer 3 Report
Although the manuscript provides interesting observations on the physiological responses of soybean to exogenous application of ABA, it is required significant improvement to be considered for publication. The authors should pay more attention in presentation of data and description of information.
The way of presenting the data lacks quality to be published. The following are some suggestions regarding this point.
The authors did not report dispersion of the mean responses in Table 1 and Fig. 1. Use SD or SE to show variability of measured responses.
Use a consistent labeling throughout the manuscript. For example, closed symbols indicate +ABA in Figs. 1 and 6, but closed symbols indicate -ABA in Figs. 4, 5, and 7. This could potentially make difficult to read through the text.
Put statistical significance codes in Figs. 8 and 9. The post-hoc analysis results from the 2-way ANOVA can be reported using different letters like in Fig. 6.
Keep in mind that the journal format requires the Materials and Methods section follows the other sections. This means information about some variables like the full names of Pn, gs, Tr, etc. should be given in the Results section first. Also, declare the exogenous ABA was applied to the soil, not to the plants, earlier in the text. Perhaps in the Introduction section. I did not notice that the ABA was treated as a soil drench until mentioned in the Materials and Methods section. Some researchers treat exogenous ABA through foliar application, so clarification of the ABA treatment early in the text will help readers understand why the authors measured soil related parameters and match these with the leaf physiology parameters.
I left minor concerns in the attached file.
Comments for author File: 
Comments.pdf
Author Response
Response to Reviewer #3
1. Although the manuscript provides interesting observations on the physiological responses of soybean to exogenous application of ABA, it is required significant improvement to be considered for publication. The authors should pay more attention in presentation of data and description of information.
Authors’ response: We are so grateful to the reviewer for this comment and while we re-written some parts of the manuscript we focused on presenting the data in a clearer and direct fashion (Please see the revised results and discussion)
2. The authors did not report dispersion of the mean responses in Table 1 and Fig. 1. Use SD or SE to show variability of measured responses.
Authors’ response: We agree with the comment of this reviewer and in the revised version we have inserted in Table 1 and Fig. 1 the SE of the mean to show variability of the measured responses.
3.Use a consistent labeling throughout the manuscript. For example, closed symbols indicate +ABA in Figs. 1 and 6, but closed symbols indicate -ABA in Figs. 4, 5, and 7. This could potentially make difficult to read through the text.
Authors’ response: We have followed the advice of this reviewer and in the revised version we have used consistently symbols to indicate +ABA and -ABA in each figure.
4. Put statistical significance codes in Figs. 8 and 9. The post-hoc analysis results from the 2-way ANOVA can be reported using different letters like in Fig. 6.
Authors’ response: We have followed the advice of this reviewer and in the revised version we have inserted statistical significance codes in Figs. 8 and 9.
5. Keep in mind that the journal format requires the Materials and Methods section follows the other sections. This means information about some variables like the full names of Pn, gs, Tr, etc. should be given in the Results section first. Also, declare the exogenous ABA was applied to the soil, not to the plants, earlier in the text. Perhaps in the Introduction section. I did not notice that the ABA was treated as a soil drench until mentioned in the Materials and Methods section. Some researchers treat exogenous ABA through foliar application, so clarification of the ABA treatment early in the text will help readers understand why the authors measured soil related parameters and match these with the leaf physiology parameters.
Authors’ response: We are so grateful to the reviewer for these comments and in the revised version we have added the full name of Pn, gs, Tr, FC, WUEi etc. We had added the methods to applied ABA in Line 17 in the abstract and L68-69 in the introduction.
6.I left minor concerns in the attached file.
Authors’ response: We agree with the minor comments of this reviewer and in the revised version we have attended each of them according to the suggestions of the reviewer (they are all highlight in blue).
Author Response File: Author Response.docx
Reviewer 4 Report
The manuscript looks interesting and presents a novel research on ABA. The authors need to address the following comments -
1. The Y axes in all figures (graphs) need to include major and minor ticks
2. Fig.1 - Please include the error bars
3. Fig.6 - Image needs to be inverted.
4. What analysis was performed by the ELISA? Which figure represents the ELISA results?
Author Response
Response to Reviewer #4
1.The Y axes in all figures (graphs) need to include major and minor ticks
Authors’ response: We thank the reviewer for this comment, but if both major and minor ticks are included the Y axes get too packed. We prefer using major ticks only for more clarity.
2. Fig.1 - Please include the error bars
Authors’ response: We have followed the advice of this reviewer and have included the error bars in the revised Fig. 1.
3.Fig.6 - Image needs to be inverted.
Authors’ response: In the revised version we have rotated Fig. 6.
4. What analysis was performed by the ELISA? Which figure represents the ELISA results?
Authors’ response: We have clarified in the revised version (Materials & Methods) that The ELISA assay was used to determine the leaf ABA concentration.
Author Response File: Author Response.docx
