Genetic Variation and Phylogeny of Wabisuke Camellias by Amplified Fragment Length Polymorphism (AFLP) Analysis

Round 1

Reviewer 1 Report

Check English text and correct grammar mistakes

Barley is the name of cereal crop, Hordeum- is the genus name. Better to put 2 articles in one referense

It would be nice to have a representative sample set of related species

too long sentences, hard to understand -  line :38,56-58,157-159

line 63-64 – barley and Hordeum- put 2 article in one play

line 137 -yield change to reveal or identify, 150 –generate – obtain, 227  -Fig 1  replace –spot

for the future work - the number of related species accessions should be more than 1.

The title of the article corresponds to the content. The conclusions are confirmed by the results. The work is interesting. Results are discussed using obtained by other methods data.

To work with old tree material very difficult. This information is important. 

Author Response

Thank you for your suggestions for improving the submitted manuscript. I am sending the replies to the comments as attached word file.

I hope these are satisfactory answers.

Sincerely yours,

Yukio OZAKI

Author Response File: Author Response.docx

Reviewer 2 Report

Review of the manuscript; “Genetic Variation and Phylogeny of Wabisuke Camellias by AFLP analysis.”

The manuscript presents brief communication of the result considering the analysis of Camelia sp. using genetic distance evaluation.

It gives the insight into the genetic structure of the species origin in Japan and China. Analysis performed in the presented study are correctly designed and performed. The plant evaluation using molecular markers gives valuable information in this field. Nevertheless the data will become more sufficient with the presentation of the phenotypic data (in. eg. regarding the biocompositon) of the analyzed species. Author may think about such an challenge in the next experiment

Line 36: It is better to start the sentence with the author name instead using citation/reference number. The sentence must be change.

After line 61 in my opinion should appear short sentence considering the Camellia genome size (one of the tea accession Camellia sinensis var. sinensis is 2x=30) and direction of genome analysis (see paper of Wai et al. 2018).

The sentence regarding, - why it is important from the agronomy point of view in e.g. It is important to cultivate due to its reach flavor and health promoting function.

In the methods it would be good to underline the Philips settings and scale – in the results 1.275  max and 0,1083 min. – what is means the distance in % ?. This may clarify the results from the table 2. showing distance range. 

Line 248 (Discussion) In my opinion it would be good to change the sentence fragment – ‘have been established from……..  ‘.

Author Response

Thank you for your suggestions for improving the submitted manuscript. I am sending the replies to the comments as attached word file.

I hope these are satisfactory answers.

Sincerely yours,

Yukio OZAKI

Author Response File: Author Response.docx

Reviewer 3 Report

This is an interesting piece of research on the phylogeny of the Wabisuke Camelia showing light over their possible origins. It is intriguing, though, why the authors chose to use AFLP for this study when there are many new molecular genetic technique, such as microsatellites, SNPs , GWAS nowadays for studying the genetic polymorphism. AFLP are not used that often nowadays and its effectiveness for indicating phylogenetic relationships has been demonstrated long time ago and this is not innovative. Nevertheless  I believe that this work is worthy of publication following some major revisions and future clarification on some unclear statements. See below some of the more specific comments on the manuscript.

Introduction :

Provide the author name of all species when mentioned for the first time in the manuscript.

Line 35: What is the scientific name of the Tarokaja and Uruku cultivars? I think it is important to mention this information to distinguish them from Camelia japonica [author?) .

Lines 46- 50: These two sentences are very confusing. Both sentences indicate that Wabisuke cultivars have small flowers and degenerate anthers and are both established or derived from ‘Tarokaja’. So what is the difference between the two cases?

Line 57:  Give acronym of RFLP in full on first mention.

Line 62:  Give full name of AFLP and RAPD on first mention

Line 72:  “length around the trunk”  is the circumference. Replace.

Section 2.1:  It is difficult to follow. It would be best to present this information in a Table and summary in the text.

Section 2.2:  I think it best to break this section into several paragraphs. It is difficult to follow and should be better restructured

Line 101:  Where is the attached protocol mention in this line.  Do you mean the following protocol relating to how the PCR was carried out.

Line 101:  Insert paragraph break starting with PCR was performed….

Line 112: Again here an attached protocol is mentioned. Where is it?  Or do you mean what follows

Line 122: Best to insert a new paragraph here beginning Digital profiles …

Results

Line 138-139: can you clarify that no polymorphism was detected between the two ‘Uraku’ accessions, and not among all accessions tested in this study?

DISCUSSION

Line 228: Change “it had been appeared” with “ It has been suggested that… Or It has been postulated that…”

Line 230- 231- Can you clarify that the study confirmed in the suggested distinction between the Tarokaja and C. japonica the same classification in the two groups as previously thought? It is not too clearly expressed.  But this becomes clear at the end of the paragraph.

Line 249 to 251: Can you clarify what you mean by involvement? This is not clear. Is it suggested that these species are suggested progenitors of the C. wabisuke

Can you clarify whether C. sinensis different from the other Chinese camellias ,as it is mentioned separately?

Line 263-264: Is this argument “The results of this experiment showed that the genetic distance between wabisuke cultivars and Chinese camellias and between wabisuke cultivars and C. sinensis” not the same? That is Chinese camelia include C. sinensis?

Line 266- What is the NJ method?

Line 269- You tend to use Uraku and Tarokaja interchangeably. It is best to use one or the other to make it more consistent. I understand from your article that they are synomynous.

Line 268-269: You are here refuting that the The Tarokaja (Uraku) is derived from F1 F1 hybrid between C. pitardii and C. japonica, as proposed by Tanaka et al. Can you elaborate the evidence for this claim here based on your study.

It will help reader if you include an overall conclusion

Author Response

Thank you for your suggestions for improving the submitted manuscript. I am sending the replies to the comments as attached word file.

I hope these are satisfactory answers.

Sincerely yours,

Yukio OZAKI

Author Response File: Author Response.docx

Round 2

Reviewer 3 Report

This is a much improved version and authors have addressed some of the earlier recommendations. However, there are a few comments made in the first review that have not been addressed (see below) and it is recommended that authors be asked to consider these before acceptance for publication.

Line 35- authors have not included the scientific names of Tarokaja and Uruku cultivars as previously recommended in the first review

Lines 52-55 “In one case, only those cultivars with small flowers and degenerated anthers that are progeny of ‘Tarokaja’ are called (narrow sense) wabisuke cultivars. In other case, those cultivars with small flowers and degenerated anthers, established from ‘Tarokaja’ or C. japonica, are called (broad sense) wabisuke cultivars.” In the preview we asked to clarify the difference between the ‘narrow sense’ and ‘broad sense’ wabisuke cultivars. The difference as reported here is not clear.

Section 2.1:  We reported that this section was difficult to follow and best to present this information in a Table and summary in the text. But this has not been addressed in the revised version.

Line 125: I would remove the word ‘attached’ and rephrased this line as “…DNA product following the Applied Biosystems protocol.” 

Line 136- as above remove the word ‘attached’ and rephrased this line as “…following the Applied Biosystems protocol.”

Lines: 325-327: The comment we made in previous review has not been addressed”  Can you clarify what you mean by involvement? This is not clear. Is it suggested that these species are suggested progenitors of the C. wabisuke . Can you clarify whether C. sinensis different from the other Chinese camellias ,as it is mentioned separately?

Line 342- as in previous review “What does Nj stands for?”

Lines 344- 348 – authors did not address the comments in the previous review.  Also no conclusion was provided.

Author Response

Thank you for your helpful advice and suggestions.

I have changed the sentences as you suggested, and cultivars-listed-Table has been created for the understanding of readers.

I hope these revisions are suitable for the journal.

Sincerely yours,

Yukio OZAKI

Author Response File: Author Response.docx