Genome-Wide Distribution of Novel Ta-3A1 Mini-Satellite Repeats and Its Use for Chromosome Identification in Wheat and Related Species
Round 1
Reviewer 1 Report
The present study by Lang et al., reports on detailed analysis of physical distribution, copy number and evolutionary aspects within the bread wheat genome of the recently developed oligo probe for mini-satellite repetitive sequence Ta-3A1. It also reports on hybridization patterns of this probe across an array of close related species together with the analysis of genetic divergence of Ta-3A1 sequence across the analysed species. The results reported have importance for evolutionary studies on the wheat genome and for characterisation of novel wheat-alien germplasm used to transfer useful traits from alien species into wheat genotypes.
The manuscript is written well and the literature review is up to date. However, one general flaw should be overcome in order to improve the quality: the authors should highlight better the novelty of their analysis on the Ta-3A1 with respect to what already published on the same probe, as in some parts, the results seem similar to the results published elsewhere (see below for details). They should more clearly describe what is different here. In several cases, it is about rephrasing and detailing more.
Other comments:
Title - you should specify the name of the micro-satellite analysed, the present work is not generic, for example: "Genome-wide distribution of novel Ta-3A1 mini-satellite and its use for chromosome identification in wheat and related species"
Line 30 - Check your reference [1], there seems to be two papers listed
Line 46 – the percentage of repetitive sequences is >85% as reported by the IWGSC (your reference [25])
Lines 62-65 - I suggest here a description of the probe and the story behind (what is known so far on it from previous work, including the reason why you have chosen this probe to further analyse. Moreover, in n the work by Lang et al., 2018 (Planta, https://doi.org/10.1007/s00425-018-3033-4), this probe was already validated by ND-FISH, therefore, highlight better where is the novelty here vs. your previous work. Please refer as well to the method of TR identification.
Line 67 - Make a table with detailed and exhaustive list of all the species analysed and include their genome formula, ploidy level and the origin
Line 116 – the reference Lang et al. should be replaced with [68]?
Line 122 – not clear “in those above chromosomes”, please rephrase
Line 131 - replace the second "copy number" with "species" in the first row
Line 132-133 - this should be a part of the Figure 1 caption
Lines140-141 - which data that allow you to say this was the principal predicted mini-satellite? With what are you comparing your results?
Line 143 - replace "7D" with "7A"
Lines 146-147 – include this in the Figure 2 caption
Lines 168-169 – see the previous comment
Line 194 - panel c of the Figure 4 is very similar to the photo published in Lang et al., 2018 ([29]):here, the figure has a better resolution, yet the rest of the information is identical to what previously published - I suggest adding her some novel information that result from the present work
Line 198 - I believe it is more appropriate "to shed light" or similar, instead of "resolve"
Line 214 - do you mean bread wheat in “wheat lines”? If data is not known, then, how you can say "in contrast to"?
Line 220 - indicate the respective colour of the probes Oligo-pSc119.2 and pTa535
Lines 225-226: how this relates to your work here?
Lines 226-228: is this the results from the cited references in previous phrase or the result of the present? If this was already done, you should discuss it in the Discussion section and indicate it in the Introduction (last paragraph, where the oligo-3A1 is described)
Lines 263-264 - where is this described?
Lines 307-308: If this sentence was true, your present paper would not have much of originality, please rephrase it i.e. highlight the novelty of your work
Lines 309-312: develop better the final remarks on the possibilities for further use your new data, including the examples you refer to in these lines.
Author Response
Response to Reviewer 1 Comments
The present study by Lang et al., reports on detailed analysis of physical distribution, copy number and evolutionary aspects within the bread wheat genome of the recently developed oligo probe for mini-satellite repetitive sequence Ta-3A1. It also reports on hybridization patterns of this probe across an array of close related species together with the analysis of genetic divergence of Ta-3A1 sequence across the analysed species. The results reported have importance for evolutionary studies on the wheat genome and for characterisation of novel wheat-alien germplasm used to transfer useful traits from alien species into wheat genotypes.
Point 1: The manuscript is written well and the literature review is up to date. However, one general flaw should be overcome in order to improve the quality: the authors should highlight better the novelty of their analysis on the Ta-3A1 with respect to what already published on the same probe, as in some parts, the results seem similar to the results published elsewhere (see below for details). They should more clearly describe what is different here. In several cases, it is about rephrasing and detailing more.
Response: I agree to rephrase the descriptions for the study. Our previous study mentioned the ND-FISH probe of Ta-3A1 only in wheat. In this present study, we revealed the genome-wide physical location, sequences variation and evolutionary aspects of Ta-3A1. Also the comparative ND-FISH patterns of Ta-3A1 in different wheat related species were made.
Point 2: Other comments:
Title - you should specify the name of the micro-satellite analysed, the present work is not generic, for example: "Genome-wide distribution of novel Ta-3A1 mini-satellite and its use for chromosome identification in wheat and related species"
Response: I agree to the revision for the title as suggested.
Line 30 - Check your reference [1], there seems to be two papers listed
Response: I agree to the correct the typing error for the reference.
Line 46 – the percentage of repetitive sequences is >85% as reported by the IWGSC (your reference [25])
Response: I agree to change it as >85%.
Lines 62-65 - I suggest here a description of the probe and the story behind (what is known so far on it from previous work, including the reason why you have chosen this probe to further analyse. Moreover, in the work by Lang et al., 2018 (Planta, https://doi.org/10.1007/s00425-018-3033-4), this probe was already validated by ND-FISH, therefore, highlight better where is the novelty here vs. your previous work. Please refer as well to the method of TR identification.
Response: Our previous study mentioned the ND-FISH probe of Ta-3A1 only in wheat. In this present study, the genome-wide physical location, sequences variation, and phylogenetic analysis of Ta-3A1 were generated.
Line 67 - Make a table with detailed and exhaustive list of all the species analysed and include their genome formula, ploidy level and the origin
Response: I agree to add the detailed materials in supplementary table S1.
Line 116 – the reference Lang et al. should be replaced with [68]?
Response: I agree to change the reference of [29].
Line 122 – not clear “in those above chromosomes”, please rephrase
Response: I agree to change it to five chromosomes.
Line 131 - replace the second "copy number" with "species" in the first row
Response: I agree to make the corrections.
Line 132-133 - this should be a part of the Figure 1 caption
Response: I agree to change.
Lines140-141 - which data that allow you to say this was the principal predicted mini-satellite? With what are you comparing your results?
Response: I agree to add the compared reference of [29].
Line 143 - replace "7D" with "7A"
Response: I agree to change.
Lines 146-147 – include this in the Figure 2 caption
Response: I agree to add the change.
Lines 168-169 – see the previous comment
Line 194 - panel c of the Figure 4 is very similar to the photo published in Lang et al., 2018 ([29]):here, the figure has a better resolution, yet the rest of the information is identical to what previously published - I suggest adding her some novel information that result from the present work
Response: I agree to add the change.
Line 198 - I believe it is more appropriate "to shed light" or similar, instead of "resolve"
Response: I agree to the change.
Line 214 - do you mean bread wheat in “wheat lines”? If data is not known, then, how you can say "in contrast to"?
Response: I agree to delete the sentences.
Line 220 - indicate the respective colour of the probes Oligo-pSc119.2 and pTa535
Response: The color Oligo-pSc119.2 (green) and pTa535 (red) were added;
Lines 225-226: how this relates to your work here?
Response: The identification of individual chromosomes from Secale, Dasypyrum and Thinopyrum was determined by Oligo-pSc119.2 and Oligo-pTa535, we can follow the identification to recognize the location of Ta-3A1.
Lines 226-228: is this the results from the cited references in previous phrase or the result of the present? If this was already done, you should discuss it in the Discussion section and indicate it in the Introduction (last paragraph, where the oligo-3A1 is described)
Response: We discussed in last paragraph.
Lines 263-264 - where is this described?
Response: The web serve of B2DSC was added.
Lines 307-308: If this sentence was true, your present paper would not have much of originality, please rephrase it i.e. highlight the novelty of your work.
Response: The previous studies used the FISH signals of Oligo-3A1 probe on 3AL. For example, it is hard to determine the arms of 7A by conventional C-banding and FISH. The FISH hybridization by Ta-3A1 can be used to distinguish between the long arm and short arm of 7A.
Lines 309-312: develop better the final remarks on the possibilities for further use your new data, including the examples you refer to in these lines.
Response: I agree to edit the discussions on the point. The FISH specific hybridization sites by Ta-3A1 on the Secale, Dasypyrum and Thinopyrum chromosomes can be used to trace the specific chromatin in wheat-alien introgression lines.
Reviewer 2 Report
The authors have used sequence analysis to find a TR sequence that is widely distributed in the wheat genome and its relatives. The distribution of Ta-3A1 was then confirmed by FISH in wheat and other related species to study copy number and its distribution. The authors claim that the Ta-3A1 sequence can assist in the precise identification and documentation of novel wheat germplasm engineered by chromosome manipulation.
The authors have done an extensive study using this oligo sequence in wheat and other relatives. I have a few minor suggestions that will improve the quality of the manuscript.
In Table 1 please correct the headings and swap copy number with chromosome and change copy number to species in the second half of the table.
On line 143 it says chromosome 7D but in Fig 2c the chromosome region displayed is 7A.
Also in Fig 2 the arrows seem to be green rather than blue as written in the legend.
Please describe what Figure 4 parts a b and c are in the main text and in the legend.
In line 208 it should be table3 not 2.
Line 270 somewhat should be somehow.
Author Response
Response to Reviewer 2 Comments
The authors have used sequence analysis to find a TR sequence that is widely distributed in the wheat genome and its relatives. The distribution of Ta-3A1 was then confirmed by FISH in wheat and other related species to study copy number and its distribution. The authors claim that the Ta-3A1 sequence can assist in the precise identification and documentation of novel wheat germplasm engineered by chromosome manipulation.
The authors have done an extensive study using this oligo sequence in wheat and other relatives. I have a few minor suggestions that will improve the quality of the manuscript.
Point 1: In Table 1 please correct the headings and swap copy number with chromosome and change copy number to species in the second half of the table.
Response: I agree to the correct the typing error.
Point 2: On line 143 it says chromosome 7D but in Fig 2c the chromosome region displayed is 7A.
Response: I agree to the change 7D to 7A.
Point 3:Also in Fig 2 the arrows seem to be green rather than blue as written in the legend.
Response: I agree to the change blue to green in Fig 2.
Point 4: Please describe what Figure 4 parts a b and c are in the main text and in the legend.
Response: I agree to the change.
Point 5: In line 208 it should be table3 not 2.
Response: I agree to the change.
Point 6: Line 270 somewhat should be somehow.
Response: I agree to the change as somehow.
