Predicting Sex-Related Transcripts in the Chinese Giant Salamander (Andrias davidianus): A Transcriptomics Study, Selection Gender for Preservation, Breeding and Reintroduction

Round 1

Reviewer 1 Report

Comments for author File: Comments.pdf

Author Response

Chinese Giant salamander (Andrias davidianus) is an endangered species and classified into China National Class II’s key protected aquatic wildlife. The authors conducted gene expression studies for gonads, muscles and skins in males and females of this species. The authors found differentially expressed genes between male and female organs. This paper is acceptable as a simple resource paper. However, needs large revision the manuscript

The purpose of this study was unclear for me. The authors claimed thatdevelopment of a method for accurate sex identification is critical for successful captivebreeding. However, you have not discussed how to use DEGs data for actual sexidentification in breeding. In the first place, I’m not sure that comparison of expressedgenes is an effective approach for sex identification. Gene expression in muscles andskins might fluctuate through developmental stages, ages and seasons. Of course, largenumber of DEGs were identified between testes and ovaries. However, these tissuesshould not be available for sex identification of animals used for breeding. On the otherhand, muscles and skins are available for RNA extraction in animals used for breedingas salamanders generally have regeneration capabilities for some organs. Thus, Mspand ZP3, DEGs in muscles, are possibly good candidate genes for sex identification. Istrongly recommend you to conduct simple PCR tests for gene expression of Msp inmuscles from multiple known-sex individuals, e.g. ten males and ten females.

The authors discussed sex determination using DEGs between testes and ovaries. This is somewhat irrelevant discussion as these tissues were collected from six-year old animals. Their sexes were probably determined much early developmental stages. English was not clearly written and there were many grammatical and typing errors in the manuscript. Many sentences did not represent correctly the results and the authors’ thinking. I strongly recommend you to ask English proofreading to native speaker who has a sufficient knowledge for biological science

Response: Thank you for your important comments. We are really sorry for not presenting a clear purpose in the introduction. In the revised manuscript, we have added/changed some sentences (marked in yellow) to make it clearer. It is true that the comparison of expressed genes is not an effective approach for sex identification as many studies have strongly proved that DNA is the simple and effective approach in identification.

We agree with reviewer’ point that the MSP expression should be performed in muscles in multiple sex-known individuals if the purpose of this study is to explore the sex identified genes. However, with the reasonable revision of the purpose in our study (was to explore the sex development/hallmarks of genes), we decide not to provide these data. Of course, we have no doubt in the correctness of the reviewer's opinion.

It is true as the reviewer suggested that the sex determination is irrelevant with our purpose of this study, we thus delete this kind of description in the revised manuscript.

We have sent our manuscript to native speaker to improve the manuscript grammar and language.

Reviewer 2 Report

The manuscript of Zhao et al. entitled “Predicting sex-related transcripts in the Chinese Giant Salamander (Andrias davidianus): A transcriptomics study,  selection gender for preservation, breeding and reintroduction” describes making a reference list of genes that are highly expressed in testes and ovaries of Chinese giant salamander. The identified genes are not basically different from those in other amphibians as well as other vertebrates. However, they would be applicable for future analyses to understand the mechanisms of sex determination and sex differentiation in these curious and precious animals. On the other hand, in the abstract and introduction, the authors mention the aim for utility of these gene list for sexing, but already the very useful sex-linked DNA markers have been identified in this species by other researchers (Hu et al., 2021) and in fact they are actually useful and applied in the field work. So, the purpose to find sex-specific markers is poor in the impact. In addition, transcriptome list is hard to be applied to sexing because RNA and cDNA has to be extracted and synthesized for sexing. It is not actually useful.

 

Minor points: two words at several sentences are arranged with no space next to next. Please check them.

Author Response

The manuscript of Zhao et al. entitled “Predicting sex-related transcripts in the Chinese Giant Salamander (Andrias davidianus): A transcriptomics study, selection gender for preservation, breeding and reintroduction” describes making a reference list of genes that are highly expressed in testes and ovaries of Chinese giant salamander. The identified genes are not basically different from those in other amphibians as well as other vertebrates. However, they would be applicable for future analyses to understand the mechanisms of sex determination and sex differentiation in these curious and precious animals. On the other hand, in the abstract and introduction, the authors mention the aim for utility of these gene list for sexing, but already the very useful sex-linked DNA markers have been identified in this species by other researchers (Hu et al., 2021) and in fact they are actually useful and applied in the field work. So, the purpose to find sex-specific markers is poor in the impact. In addition, transcriptome list is hard to be applied to sexing because RNA and cDNA has to be extracted and synthesized for sexing. It is not actually useful.

Response: Thank you for your important comment. We are sorry for not expressing our purpose of this study well, and we agree with your point that our study is valuable for further study of determination and sex differentiation in this curious and precious species. We thus changed our abstract and introduction in yellow marked sentences.

Minor points: two words at several sentences are arranged with no space next to next. Please check them.

Response: Thank you for your comment. We have check these errors throughout the manuscript and corrected them accordingly.

Round 2

Reviewer 1 Report

 

Comments to authors:

 

My major point for the first manuscript was discrepancy between the purpose of this research and approaches which you applied to accomplish it. I suggested two options for revision. One is modifying the purpose to fit approaches, results and discussion. Another one is carrying out further researches to complete the purpose. The authors have selected the former, simpler way compared to the latter. I do not deny your choice. I have no more comments for revision.

Author Response

My major point for the first manuscript was discrepancy between the purpose of this research and approaches which you applied to accomplish it. I suggested two options for revision. One is modifying the purpose to fit approaches, results and discussion. Another one is carrying out further researches to complete the purpose. The authors have selected the former, simpler way compared to the latter. I do not deny your choice. I have no more comments for revision.

Response: Thank you again for your important comments. We are glad to hear that our revision could be approved by the reviewer. And, we do believe our current manuscript has been improved a lot after revision based on your suggestions.