Transcriptomic Analysis of Rice Varieties Under System of Rice Intensification (SRI) Management

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The research topic is highly meaningful, employing representative indica and japonica varieties of paddy and upland rice as experimental materials and applying AWD treatment throughout the entire growth period to analyze DEGs among different rice varieties. The writing is also well-executed. However, there are two issues that need to be addressed:

 

Since this study spans the entire growth cycle of rice, data on rice yield and agronomic traits under different water treatment conditions should be supplemented.

Are the growth periods of the four varieties consistent? For instance, do they flower at the same time? The different growth stage at which drought stress occurs can significantly influence rice growth and yield differently.

Any omics analysis generates a large number of DEGs. It is essential to integrate the analysis of DEGs with phenotypic data to derive meaningful insights.

Author Response

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Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

The current paper aimed to use RNA sequencing to identify the differentially expressed genes in upland and lowland rice under different cultivation methods.

Authors made a good molecular analysis of gene expressions after adaptation from bulk RNA sample, ea cell type and position specificity.

Therefore data interpretation and conclusions require significant re-evaluation.

Authors used DEG for conclusion. However, these methods have a two major problems:

1. The analysis was done after 50 days, when total metabolism became different between treatments. The link between causality and chain of reaction is missing, including cell type. The correct name is adaptive changes in gene expressions after treatments.
2. “Butterfly effect” = fold of changes in gene expression not equal to “value” of these genes in adaptation process. Some genes can chnege mRNA level on 10-20%, but this caused changes of downstream genes 2-3 times.

 

The results are interesting and good, but still require new interpretations.

Some details:

Line 91: “enhanced reactive oxygen scavenging activity” – may be first increased ROS accumulation after induction of imbalance between different cell types?

Line 93: “tolerance; this response” – tolerance (adaptation) and response is not the same. Response is immediate,  adaptation is result of reprogram (adaptation).

Line 95: “Reactive oxygen species (ROS) also increase during abiotic stress that cause oxidative damage to several biological molecules” – ROS production? Accumulation? ROS (H2O2 etc) can not increase itself. Moreover, the most important is a treason: more production? Less scavenging? And: damage is the last step under very high ROS local level in specific cell type. ROS play role as signaling in cell to cell communication, wheat is the most “dangerous” stress effect. https://doi.org/10.1111/tpj.16118

Line 156: “Transcriptome responses of roots to water management” – transcriptome can not respond. Moreover, you did not study response, it was adaptation.

Line 103: “The key stress-responsive genes in rice (Oryza sativa) play crucial roles in enhancing the plant's resilience to various abiotic stresses” ? It looks like author confuse response and adaptation.

Fig 2- very good illustration the differences between leaf and root in structure and response to growth conditions. Very well shown possible differences between cell type ratio in root and leaf: leaf keep similar cell type, while root may have different cell type raio because eof more flexibility in structure/phenotypes.

Lines 212- 220: it will be great to show at least briefly location: cell cycle in the main root, or side root? ROS accumulation/scavenging in which cell? If treatemts in your case gave you more side-root, then expression of cell cycle gene is readout of new local hormonal balance, which can not be detect by transcriptome since you study bulk RNA and miss local information.

Similar comments for other concussions: maybe different mRNA samples have different cell type ratio, and this is lead to different DEG. This is another problem with DEG meaning after long-term treatments.

Line 550: “50 DAT” ?? – it is not a response, it is adaptation.

Author Response

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Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

Summary

This paper studies gene changes (transcriptomics) in four rice types. The rice grew with two irrigation methods: normal flooding and alternate wetting and drying (SRI). The authors examined gene changes in roots and leaves. They wanted to see how rice adapts to different water settings. This study found each rice type and organ has unique ways to handle water stress.​

 

Major Comments

One rice type did not have enough leaf data. This makes it harder to compare all results fairly.​

The experiment was in controlled tanks (lysimeters), not in open fields. This may not show how rice reacts in real farms.​

The same fertilizers were used for both groups. The authors should make sure these were really equal and say if any differences may affect results.​

The paper did not measure how much rice was produced. The gene changes are interesting, but we do not know if they help real crop yield or plant growth.​

The methods for choosing which genes were important should be explained better. The authors need to say why these numbers were used.​

Please deposit all raw sequencing data in a public database such as the NCBI Sequence Read Archive (SRA). After submission, provide the SRA accession numbers in the manuscript.​​

 

Minor Comments

Some tables and figures are hard to read. Use bigger fonts or clearer labels.​

Some explanations are long or repeat ideas. Make the writing shorter and simpler when possible.​

Define all special words and keep language easy to follow.​

Comments on the Quality of English Language

The English could be improved to more clearly express the research.

Author Response

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Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

It is a pity that no supplement the data on yield and agronomic traits, which would have made the analysis more meaningful. It is hoped that future work can be systematically conducted in conjunction with practical production, rather than being limited only to the analysis of gene expression.

Author Response

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Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

Thank you very much for a very professional response. 

Minor points still need.

Line 654: I would suggest to add also cell-type specific dynamic of adapation = sevarl time points, starting from the early response.

Please, also consider that single cell RNA seq and spaitail transcriptomic can nit solve a problem: the problem is cell to cell communication which can be studied only in situ with 3D analysis = spatial cell/molecular biology platform.

My best regards!

Author Response

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Reviewer 3 Report

Comments and Suggestions for Authors

I have checked the revised manuscript.

Authors have diligently carried out revisions based on the reviewer's comments.

I recommend the manuscript for publication in its current form. 

Comments on the Quality of English Language

The English is fine.

Author Response

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Author Response File: Author Response.pdf