Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: A SIMPLE AND EFFICIENT CLONING SYSTEM FOR CRISPR/CAS9-MEDIATED GENOME EDITING IN RICE
Review round: 2
Reviewer: 1

Output:
Basic reporting: Technology involving the targeted mutagenesis of plants using programmable nucleases has been developing rapidly and has enormous potential in next-generation plant breeding. Notably, the clustered regularly interspaced short palindromic repeats/Cas9 (CRISPR/Cas9) system has paved the way for the development of rapid and cost-effective procedures to create new mutant populations in plants. Genome-edited plants from multiple species have been produced successfully using a method in which a Cas9-guide RNA (gRNA) expression cassette and selectable marker are integrated into the genomic DNA via Agrobacterium tumefaciens-mediated transformation or particle bombardment. Although CRISPR/Cas9 integration increases the chance of off-target modifications and foreign DNA sequences cause legislative concerns about genetically modified organisms, transformation of plants with Cas9-guide RNA (gRNA) expression cassette DNA is most popular and widely used approach for making genome-edited plants.

In the manuscript (peerj-42378), the authors established simple and efficient system for constructing CRISPR/Cas9-gRNA expression vectors to produce genome-edited rice plants. In this system, overlapping PCR and Gateway/Golden Gate system were successfully used for reducing the step and time for vector construction. The reviewer considers that present CRISPR/Cas9-gRNA expression vector construction system is highly useful to reduce molecular lab works such as PCR, subcloning, ligation. Although figures for the former version of the manuscript were too preliminary to be published in the The PeerJ, the figures and descriptions for these figures were largely improved in the present revised manuscript. Therefore, the manuscript will be acceptable for the publication in The PeerJ.
Experimental design: OK
Validity of the findings: OK
Additional comments: All concerns indicated by the reviewer indicated were correctly revised in the manuscript. Therefore, the reviewer thinks that the manuscript is now suitable for publication in The PeerJ.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: THE GLOBAL MUSEUM: NATURAL HISTORY COLLECTIONS AND THE FUTURE OF EVOLUTIONARY SCIENCE AND PUBLIC EDUCATION
Review round: 1
Reviewer: 1

Output:
Basic reporting: Manuscripts like this with multiple authors contributing are always difficult to assemble and to assess.  There is plenty of value in this paper, but the challenge comes from integration and then organization and I feel like the manuscript suffers as a result (which is not unusual of such efforts).  I also represent a reviewer who may be overly opiniated about certain aspects that may not be important to the authors or to readers.  It is hard to assess exactly who the audience for this manuscript is (and maybe this should be discussed with even more clarity at the beginning).  I would say currently, it is the broader scientific community, the educated public and the museum community in different ways throughout.  That is ambitious and problematic.  I would argue different sections target these different audiences and assumptions are made throughout about what readers are going to be familiar with, when they may not be.  In addition, specific examples (e.g., the collecting example of the kingfisher) are not presented with enough context to be clear who the audience actually is.  This can be unproductive.  Below, I found myself pulling sentences out throughout the manuscript where I felt the authors were stating things in unproductive ways or straying away from what they were trying to say, or simply weren’t clear enough for the average reader to understand what they were talking about.

The biggest issue to me is that, as currently written, the “Global Collection” they envision isn’t clearly enough defined in terms of the actual human capacity and expertise needed to carry it off.  This extends to the public education aspect as well.  I also do not see enough of a “plan” to actually develop this internationally.  There are glimmers of this here and there, but that is all and I think this is an opportunity argue for making this effort truly global.

I also think anthropocentricism is a trap we all fall into.  We are not and should not be doing biodiversity science and collections-based research only for humans.

I think the issue of fossil material needs to be developed more here.  Again, there are glimmers in places, but just that.  Museums are the place where this material is archived.
Experimental design: No comment
Validity of the findings: See some specific comments in the general comments section below.
Additional comments: My comments come after quoted sentences below:

“and even provide solutions to climate change, global health and food security challenges.”

Statements written like this to me undervalue the role of these collections.

“In this perspective, we discuss challenges to the realization of the full potential of natural history collections and museums to serve society.”

If the collections community spends all its efforts emphasizing the value of biodiversity to humans, the planet is probably doomed.

“Through extensive exhibits and public programming and by hosting rich communities of amateurs, students, and researchers at all stages of their careers, they provide a place-based window to focus on integration of science and discovery, as well as a locus for community engagement.”

This is what could be, but it also illustrates the issues.  Most museums cannot have extensive exhibits.  Many do not host rich communities of people, they could, but most of these communities are small.

“These new directions include the possibility to ask new, often interdisciplinary questions in basic and applied science; inform biomimetic design; and even provide solutions to climate change, global health and food security challenges.”

Sure, but they also serve the local community and monitor locally.

“We also explore ways in which improved infrastructure will allow higher quality science and increased opportunities for interdisciplinary research and communication, as well as new uses of collections.”

Infrastructure needs expertise

“Natural history museums, which harbour extensive records of biological diversity, have always
been meeting places for scientists, amateurs, and the public.”

They harbor THE basic authoritative records of biological diversity (observations are useful, but essentially derivative data.

“As such, museums are still considered trusted resources, at a time when many other institutions are bitterly mistrusted (Foley, 2015).”  Does this mean they may not be trusted in the future?

“they are also actively used and curated by professional scientists to answer pressing problems in biology and beyond”
I’d argue that this isn’t necessarily true in most museum.  Hence, a major problem.

“This wealth of metadata…”  Pathogens really aren’t “metadata,” they are specimens in and of themselves.

“to global change (tracking shifts in phenotype across specimens through time; Jones & Daehler, 2018)”

While I agree with this, the truth is that local change is just as important as global change and our community needs to recognize that.

“Participants, most of whom were based at natural history museums (broadly defined to also include botanical gardens; see below)”

I would not call this necessarily “broadly defined.”  By this, I mean that collections-based institutions include seed  banks et cet.

“Additionally, citizen science increasingly contributes to collections, which today are housed all over the world, and serve as gems of diverse global centres of cutting-edge research (see Fig. 1).”

While using “Gems” has “lustre,” they are equally factories of knowledge and they are centers of education about the natural world for an increasingly urban human population. (e.g., Bates, 2007).  We always need to be on guard about communicating the true breadth of what we do.

“Natural history museums may be located at universities, sometimes without exhibits, or may include public exhibits, such as typically occurs in national, state or regional entities.”

The issue of museums and collections should probably be dealt with more directly.  They may be private/public entities.

“Regionality therefore, can be considered a strength of collections and fulfills an important role in sustaining regional pride in biodiversity.”

This an anthropocentric view that I hope we can combat someday.  While “regional pride” is an outcome, I see the goal as regional (and local) knowledge, active research and education, and appreciation of the value of biodiversity.

“Still, given the increasingly connected network of regional museums, the local depth provided by such regionality would be impossible to recover – if only for practical reasons - in a single, global museum, were it composed solely of physical specimens.

Because of the value of local monitoring, this should not even be the goal (and I know it isn’t).  There are simply different scientific and educational reasons requiring global, regional and local levels.

“A grim example is the Brazil National Museum in Rio de Janeiro, where a fire destroyed an estimated 90% of the collections in several divisions in September 2018 (Phillips, 2018).”

This is a little too brief about what was lost and what might be done to mitigate the knowledge and data loss in such cases (and to suggest that rebuilding such collections is possible, if humans would see these institutions in the same way they see Notre Dame).

“Creating redundancies in collections, especially for extant species and genetic resource collections, is key to ensuring the longevity of these samples and associated data. Initiatives such as the Global Genome Biodiversity Network (GGBN; Droege et al., 2016) aim to collect, catalogue, and “democratise” genomic resources across global collections, covering 50,626 species (as of 18 march 2019).”

This is a weird statement and it is not clear to me how it relates to the previous examples of protecting our biodiversity knowledge base from catastrophic losses of knowledge.

186 “… of now-extinct animal species, such as the Tasmanian tiger, the golden toad, and the Hawaiian crow.”

Hawaiian Crow is not extinct.  I would encourage use of scientific names throughout.

192  :These require… storage resources from those that traditionally constitute museum infrastructures, namely large scale and secure long-term storage of image data.”

This is true, but we, as a community, need to be specific, these require digital storage which is distinct from the physical, and they also require critical curation by experts.

“Genomic analyses of single bone fragments can inform on the evolutionary and demographic history of our own species (e.g., Slon et al., 2018).”

Great, but this is one in millions of species out there on the planet.

“…many collections now come from large scale ecological studies (e.g. NEON in the
US).”

I would argue it is still “some” and too often ecological research is conducted in ways that could archive specimen data, but choose not to (and of course local/regional museums often aren’t adequately staffed to effectively argue that they could assist in such efforts).

“Museum spaces ideally are filled with students who learn to think anti-disciplinarily and appreciate the importance of the specimen. These spaces can therefore be considered ‘Innovation Incubators’ where a next generation of critical thinkers in biology and beyond will be trained.”

This may be a visionary statement, but I think it misses what museums bring to the system.  They bring the expertise of taxonomic groups necessary to provide context innovation incubators too.  If we fill our museums with innovation incubators, they are likely to be a bunch of smart folks, but will that help biodiversity itself?  I wonder if a coordinated effort that includes such innovators is what is needed.

“In addition to such observations recorded in the field, however, collected specimens (when available) offer additional options for confirming or extending the original work using new analytical techniques.”

Boakes et al. 2010 is a useful comparative example for a group of birds.

“Places are not simply a semantic convenience. It is a meaningful lens for viewing the world because it is orderly with respect to geographic space.”

Change “It” to [They]

284 “Place-based learning and education is well developed (Gruenewald and Smith, 2014) and provides a context for local understanding and societal change.”

This paragraph meant little to me after reading it several times.  The following paragraph also didn’t work well for me.  “Place-based”  is being defined based on the specimen and where it comes from, I think.  While that is one version of “Place-based.”  Another if where a specimen is institutionally which allows for comparative study with specimens from other places.  Again, both have critical value for biodiversity.

Line 315.  Delete “some”

“For instance, for taxonomy, having the virtual, global, workbench of the Barcode Of Life Database BOLD (www.boldsystems.org; Ratnasingham & Hebert, 2007) allowed taxonomists globally to harmonise species delineations by collectively analyzing and interpreting DNA barcode patterns from global rather than regional data sets.”

I find this a fairly bold statement without much ancillary data in terms of publications provided.  This is not to say the effort was not valuable.  But were species delimitations really established and effectively coordinated with taxonomic efforts?  To me, Barcodes almost are more like ebird records, they are snippets of data, that while useful, really go only so far in terms of helping taxonomists.

Line 339.  This statement simply isn’t true.  Rarity has been only one criterion.  We need to stop saying things like this.  In many groups of organisms (e.g., micro-organisms), we really do not even know what rarity means.  With many vertebrates, we actually no longer collect rare taxa (which also may be a mistake in the long run, Rocha et al. 2014).

“In addition, assumptions about species ID based on morphology may be falsified by genetic data (DeSalle et al. 2005) - but also the reverse - revealing an unexpectedly high level of cryptic diversity in certain groups (e.g., Hebert et al., 2004).

Does this essentially highlight limitations associated with approaches that don’t necessarily include expertise (and other types of data)?

“Other ventures include the above-mentioned BOLD (with iBOL extending its coverage) which holds 6.6M barcode records across 0.29M species, many of which are commonly-occurring. Future collections should continue to expand with specimens sampled widely across biodiversity, but in addition should amass commonly-occurring species, which can serve as environmental monitors, especially when sufficient metadata is also collected.”

This statement is simply bizarre to me in relation to the paragraph above and I don’t think the numbers are really different.  Yes, a lot of interesting sampling has been done with barcoding.  Is 22.7 individuals per “species” really all that different than what is in specimen collections?  Are there vouchers of all these bar-coded individuals?  That would be valuable information if it is the case.

Line 362.  While this sentence is true in the context of technology, it isn’t true in a “place-based” sense.

390  “Digitization of collections will be increasingly important in this respect; there are many valuable but undigitized collections residing in museums.”

This paragraph lauds the digitzation efforts which I support completely, but fails to acknowledge the essential need to reach out and digitize collections outside of developed countries (and to prioritize hyper-diverse groups like insects and invertebrates).

Line 394.  These donations should be made not for “posterity” but for science.

Bolotov et al. (2018) could infer from freshwater pearl mussel collections that morphology has changed in time correlated to environmental alteration and climate change. Based on historical and recent specimens from extensive geographical sampling, the authors concluded that the latter may well have accelerated the population decline in pearl mussels over the last 100 years. The study underlines the importance of preserving large collections (many individuals) to enable meaningful statistical analysis of morphological measurements.

These sentences read to me like the collecting caused the declines.  This needs to be reworded

Line 413.  My guess is most readers unfamiliar with the community won’t rembmer what iDigBio is.

Line 414.  There should be citation here.  iDigBio annual reports?

417 A major question for the future is how the community should greatly expand the scope of digitized specimens.

Cite the new Biodiversity Collection Network report that looks forward specifically with respect to digitization efforts in the United States?

Line 418.  “haphazardly” is a tough word for me to read in scientific paper.  “Incidentally” would be better.
I think it fair to criticize the approach taken in the U.S. with respect to project-based digitization, but recognize that we could be shooting ourselves in the foot in such situations.  We’ve spent a lot of money bar-coding things, and this could be criticized as well.

Line 426.  But the truth in all this remains that even in well known taxonomic groups, expertise is critically needed to verify efforts to digitize data and effectively curate it.

“Knowing the limits in geographical and temporal distribution and the size of the collection, especially for the most common groups, may suffice for further scientific analysis.”

I don’t think there is enough information here for readers to understand what is being said here.  It is an important point worthy of more discussion.

“can easily be supplemented by citizen science initiatives (e.g iNaturalist),”

Only true if species identification is easily made, clearly not the case for many such taxa.

Line 461.  Add Beespotter (https://beespotter.org/)

Line 467.  …Data are…

475 An abundance of undescribed species is only the tip of the iceberg on the amount of data undiscovered and undocumented in the world’s museums.

It is fine to point this out, but why not say what the issues are to solving this problem too?

Line 481. Fix this sentence: “In UNITE, fungal species hypotheses are generated and named, but also tagged with a citable digital object identifiers (DOIs) so THEY can be unambiguously communicated, allowing harmonisation of species concepts throughout communities.

506 ?Natural history museums should continue their efforts to train scientists and artists to develop novel solutions to emerging problems, especially as we face an increasingly uncertain environmental future.?

For me, an emerging problem  identified in the paragraphs above is the lack of expertise necessary to rigorously describe diversity.

Line 509.  This paragraph, to me, doesn’t go far enough in stating that museum networks need to be global and there have to be initiatives and strategies developed to support and improve facilities throughout the world.

Line 546.  “This sad event,”  This is far too vague to be useful to most readers.  What is sad for the kingfisher? Or sad the way it played out in the media.  And let’s be realistic.  There probably are ways in which this could have been done differently.  Also, even mentioning this event give credence to a small, but vocal group of people with viewpoints that probably do not warrant validation by our community.

Line 549.  “Additionally, the public in this case did not appreciate the relative insignificance of scientific collecting as an agent of species loss as compared with habitat loss and introduced or feral predators, such as house cats.

This is overstating things in a way that doesn’t make sense to do.  Some members of the public felt this way, but most didn’t, or they hadn’t or didn’t even think about it.  Our community needs to always look for ways to effectively engage the vast majority of people who either haven’t thought about the issues or who support what we do, and this is not a minority.

Line 556.  There could be more citations here to other examples (e.g., Rocha et al. 2014)

“Discussions should be held that address how we can best leverage collecting activities across the Global Museum and that planning should lead to a global effort to more rigorously inventory biodiversity.”

This is an ongoing discussion that will never end, and that is a good thing.

Line 594.  These kinds of threads of ideas are useful but come across as disjointed in this manuscript, the blood of birds is not going to all that different from the blood of other things.  I think could either be removed or deleted.  At the very least it comes across as a weird way to end this paper on the Global museum.

Line 630.  I hope these data are used for much more than just documenting the anthropogenic influences on biodiversity.

I would argue that space in the paper associated with genomic data, also be allocated to how these data will help understand the phenotype.

The conclusion of this manuscript recognizes that taxonomic expertise is needed, but doesn’t clearly articulate what needs to be done about this.

714.  A. Zan

720.  Molecular Ecology (capitalization)


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: THE GLOBAL MUSEUM: NATURAL HISTORY COLLECTIONS AND THE FUTURE OF EVOLUTIONARY SCIENCE AND PUBLIC EDUCATION
Review round: 1
Reviewer: 2

Output:
Basic reporting: Clear, unambiguous, professional English language used throughout. YES

Intro & background to show context. Literature well referenced & relevant.  YES

Structure conforms to PeerJ standards, discipline norm, or improved for clarity. YES

Is the review of broad and cross-disciplinary interest and within the scope of the journal? YES

Has the field been reviewed recently? If so, is there a good reason for this review (different point of view, accessible to a different audience, etc.)?
To my knowledge the field has not been reviewed since all of the Systematic Agenda 2000 reports (not cited) were produced some years ago. The more recent literature is cited.

Does the Introduction adequately introduce the subject and make it clear who the
audience is/what the motivation is?
See comments
Experimental design: Most of these are not relevant
Yes, the article is relevant for Peerj
Validity of the findings: Multiple of these are not met. See comments
Additional comments: First, I want to say I liked this even though, as the authors will see, I was very frustrated at times.  It is important to keep pointing out the importance of museums, especially in a rapidly changing world.

Talking about the state of museums and the science done in them, and proposing solutions and agendas goes back decades, but the main talking points and initiatives began in the 1990’s with a series of NSF-supported workshops that also generated meetings in Europe.  These NSF-supported studies have continued over the years. They have been important for articulating the importance of collections and museums.  I would guess that, collectively, museums may be better off now then ever before.  This is not to say some are not in deeper trouble than others.  And obviously there are challenges.

Museums and their health cannot be separated from their role in pushing forward science and education and in solving societal problems.  Without those, there is little reason for society to support museums, except perhaps as pure entertainment.  This paper does not articulate this strongly enough, because it remains the framework of the future of museums.

It is not clear to me who the audience is.  Is it the general academic public rather than museum professional scientists (evolutionary biologists?) and administrators per se (and their public?).  If the former, what exactly is the major thing the authors want the audience to take away from this article, other than modern museums do a lot of things they didn’t do in the past? And that they need support?

The introduction does not mention “Global Museum” and given that it appears frequently, I wondered if this is a critical theme of the paper or simply a cute handle.  If it is a metaphor rather than a concrete idea of a plan, it should be so stated.  If it is an objective, then so state.  Right now I have no idea if it is a vision or an apparition.

line 111-118/ it might be good here to give the reader some very general stats.  Local museums versus the big ones: “increasingly held in local museums”?  If you are talking about “local” museums in London, Berlin, New York, Washington, then that may be true.  But by “local” do you mean small museums in small towns and cities?  Are collections “increasingly” held in these?
Percentage-wise that probably is not true.

And, yes citizen science is increasing, but what exactly is its impact on collections-based science, and in the growth of collections, given the multitudinous laws trying to protect biodiversity?  And the example of citizen science in table 4 is nice but seems pretty trivial as citizen science in which citizens actually do science.

And stop using “global museum” until you specifically say what that is.  Given that in some countries (US-especially through iDigBio) there are multiple “virtual” freely- accessible databases, one could call those global in some sense (at least for vertebrates). But this is much less true of other global collections, even the very big collections of Europe in some instances.  So I have no idea what “global museum” means. Or even whether it is a reasonable expectation over the long term given the enormous costs of digitization and especially verification.

170/ I applaud GBIF but even a lot of its data on specimens are highly suspect (their IDs, locality data), especially from the species-diverse regions of the world.  It is not what museums need, nor is it a model in my opinion. VertNet has much higher quality data (it goes into GBIF also).

209 et seq/  The justification for collecting is good, but not strong enough.  Public ideas about collecting are very fragmentary or even nonexistent.  There are multiple issues such that this perhaps needs its own section.  Some issues perhaps to talk about: (1) why and how collecting should be defended in the face of increasing anti-collecting voices, (2) the need for vouchers of all types of specimen data (just don’t bleed for dna, etc as some museums do regularly) because field and museum IDs are often wrong, or there is no solid evidence for the taxon at a given locality (a voucher is needed), taxonomic changes, etc). There is a literature on all this.

How do we convince the public and policymakers about the importance of collecting.  There are studies about the very low impact of collecting on populations.  There are voices out there saying collecting is driving things extinct.  Text about ethics? Permits? Laws?  Some of this is briefly mentioned.  BUT note, there are many biodiversity scientists, including evolutionary biologists, ecologists, etc, who are against collecting.  Whole countries (India). Your article should speak to them also.

lines 257 et seq.  It is no surprise that a database starts out using specimens, then allows dicey observational data, and the proportion of specimen data goes down.  Duh?  The solution is for museums to do more collecting, but that will never keep up with millions of bird watchers.  At least those millions of observations can be examined statistically for outliers.  I think here as elsewhere, the paragraph (or sections) mixex multiple topics.  Have a separate section on collecting, separate from databases.  There might be some additional self-reflection on the part of European museums as far as collecting.  My impression is that US museums do far more collecting (with vouchers) than European museums, with stricter laws, more accountability etc.  I would be exceedingly happy to be wrong on this.  At least in the US, curators, staff and students better adhere to all the laws or they can get fired.

274/ Place-based discovery
This section following on previous discussion seems a bit out of place (no pun intended).  Of course, “place-based” is the mantra for so much biodiversity/sustainability studies.  But I do not quite get what is being said here.  Evolutionary biology and collections are rarely “place-based” but at a larger scale.  Some of this seemingly refers to specimens at a local level, and education activities at a locality; there is a mix.
At the same time, collecting at local scales is critical (think of the German insect collections and their connection to populational declines). Without collections, those studies would have been impossible.
It seems to me that if one had a section about the critical importance of collections, some of this might go there.

A lack of a common thread here is shown in lines 324-326 mixing developing museums and DNA barcoding in a single sentence.
This section needs rethinking. It is not cohesive to me.

330  The global museum

I would not call or infer BOLD to be a global museum. That is the best example?  Basically a database of CO1 sequences?  And the last thing it does is “harmonize” species delineations (the 2% rule??--is this being taken as scientifically serious?).
Barcoding plays a role in various kinds of studies, yes, but it is also antithetical to museum scholarship in multiple ways. There are many many papers written on this but not cited.

If you are going to articulate a vision of a global museum, then do it.  There are some good ideas here, but what is the vision for linking data about phenomes and genomes across museums in such a way that it actually is synergistic to advancing answers to scientific questions?  That is a vision.  Online databases per se do not do that (except perhaps with distributional data).  And NEON was NEVER a museum-oriented program that would “help document current biodiversity and variation of common species across the globe.” Those are “place-based” observatories that do place-based ecology; museums were envisioned to be the depository for all the common specimens collected for other studies, and museums did not want that.
Metaphorically, perhaps think about the giant network of seismic monitors as a model of how to link museums in gathering and integrating museum data (this is perhaps a wild thought I know).  Again a “global museum” is not a linkage of specimen databases.  That is relatively simple (think vertnet).  Rather, how does one gather comparative data across taxonomic groups so that an investigator in Gothenburg can actually study specimens in Washington, D. C., from Gothenburg.

Of course, there are several billion specimens in the world, so one needs a plan.  There are instances in many museums of collections having digitized specimens (anthropology, CT scans) and there are multiple ongoing national initiatives as potential models for ideas.  And what about digitized literature in museums?  There are lots of historical museum science journals not yet digitized.

This entire section does not articulate a vision for the “global museum”.  Having local citizen science data contributes little to a “global museum” of science unless it is can be scaled globally like e-bird.  But you don’t need museums for e-bird-like projects.  Having citizen scientists collect all kinds of invertebrates (with no mention about how such rampant collecting can decimate populations—think egg-collecting in Europe, especially Britain) but not mentioning the pressures on museum staff to deal with this does not present the whole picture.

407/Further increasing…

This is a good section.  One thing not discussed is the idea that names applied to specimens will change as science progresses.  But data will remain.  We do not want names applied to specimens, as contrast to names on the original labels, to remain stagnant.  Good museums do indeed worry about synonymy, and this can be handled in databases, but it is also up to an outside investigator to know what they want to do.  The key here is to track all historical changes.  One reason why this paper could have devoted more to the promotion of taxonomy and nomenclature which are the bed-rock of information systems.

Not to mention VertNet as perhaps the best example of combining data bases and vetting them is short-sighted.

453/big clades-large collections  I found this section to be a bit removed from reality.  Envision trying to deal with an insect (or other invertebrate) collection with millions upon millions of specimens.  It has taken decades to century to curate that.  The solution to the problems is professional staff and infrastructure; then one can digitize, clear up the names, distributions and taxonomy and systematic relationships.

I really object to the notion that museum scientists can endorse DNA barcoding as a way to (line 478) “provide a good framework against which the extent of diversity deposited at natural history collections can be measured.”  This forgets everything that museums stand for and what they have done through their history.  DNA sequences are not taxa.  If a museum has digitized its collection, even with old names (which can generally be reconciled fairly easily in many groups), then that is the “point” estimate of the taxonomic diversity in that collection.  Again, this not saying that genetic data can’t be useful.  But trying, again, to “harmonize” species concepts throughout communities is saying that there are no scientific concepts involved in museum specimens and diversity, just a CO1 gene tree.  Museum collections are based on taxonomy, nomenclature, and rules, and scientific debates about what taxa are, as well as the hard work of thousands of scientists and staff.  There is little thought in this section about the history and function of museums and certainly about what collections represent. Or how they are managed.

Reference collections (491).  Surely there is a bigger vision that can be articulated here.  Shouldn’t we want genomes and phenomes to be linked to each specimen?  In the big picture?  Taxa are comprised of individuals with variability.  Surely a “metabarcode” is typological of a given taxon, a presumptive evolutionary unit.
And this paper keeps coming back to barcoding when the systematic world has effectively passed that by.  Barcoding is about identification and it has a role there. But scientists are building more and more trees based on large-scale genomic data. There are efforts, not mentioned, that will have genomes of most vertebrates in a decade or so.  This is what museum science represent in the future, along with a lot of other things.

496  It seems to me that this paragraph and others below need to be part of bigger questions that promote science in museums. And this should be a separate section.

If this paper is designed to tout the importance of museums and collections, then more expansive and concrete questions need to be addressed.

1. How does a vision of a global museum get realized? Who is part of this? What is the framework and cost of connectivity?  I would say this will be a lot of money. OK, how will this happen given different funding models and the fact that museums are facing numerous fiscal and other challenges (not all of which were articulated strongly enough)?  So how does one get policymakers at various levels, and the public, to say this is a top priority?  There is a lack of realism about the will of the public here.

2. What has to be done across the very wide spectrum of museum situations and funding sources that need to happen to make a “global museum” possible.  There needs to be more discussion about how museums can make themselves more necessary relative to other things the public can spend their money on.  Just remember, if you do not articulate a real vision, no one will follow.  In the US, astrophysicists periodically go to Congress and ask for billions (yes, billions) of dollars (for instrumentation) and they have been very successful.  They are united.  Biodiversity scientists are not.

3. Some other random thoughts. Evolutionary biology and public education are in the title, but surprisingly little is focused on these.  Politically, I do not think “evolutionary biology” will sell a Global Museum idea.  “Science” might.

I think this paper needs significant reorganization.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: THE GLOBAL MUSEUM: NATURAL HISTORY COLLECTIONS AND THE FUTURE OF EVOLUTIONARY SCIENCE AND PUBLIC EDUCATION
Review round: 1
Reviewer: 3

Output:
Basic reporting: This article is said to be a literature review, but it is much more of an opinion piece than a well-done and comprehensive literature review. It is the result of a what sounds like a very interesting workshop held in 2017, and as such is woefully out of date. The authors also make a point of stating that there were contributors from the Global South at the workshop, but none of the authors of the paper is, all are from either northern Europe or the United States, with the majority from Sweden. This is not bad per se, but rather limits the scope of opinion presented.

The manuscript is relatively well-written, but is quite repetitive in parts and there are lots of what I would call “motherhood and apple pie statements” such as stating that natural history museums are places where the public and science meet – this is hardly novel, and has been the strapline of several papers recently (2018, 2019). I am also not quite sure what the authors mean when they say museums need “sustenance” (line 93) – I thought at first they meant funding, but the word funding follows, so its unclear. In line 17 they state that European museums were established with imperial ambitions, but I would argue this is the case for US museums as well – they were largely established with the expansionist program westwards. This makes me wonder how much the authors are au fait with the history of natural history collections.  I am also not quite sure who the audience is for this paper. It alternately encourages and admonishes museums, but statements such as “Discussions should be held…”, “Museums should……” are not particularly helpful. Is the audience museums themselves – i.e., are these evolutionary biologists telling museums what they ought to be doing in the future (if so not a literature review, see below)? Or is the audience those funding and using museums, including the public, in order to garner support for these collections in the future? This ambiguity runs throughout the manuscript.

The literature referenced is the tip of the iceberg when related to the topic of museum collection use in other fields. The authors primarily cited their own work (I didn’t count, but was left with the overwhelming impression) and certainly are not providing a literature review that would let a reader explore just how museums and their collections have been used. As one of many examples, the discussion of the impact on collecting by public opprobrium (lines 540-550 approximately) is limited to a single example, while there is ample grey literature and blog content that could be used and referenced here. The manuscript is broad, but with the audience not clearly defined I am not sure of its ultimate impact.
Experimental design: Content is within aims and scope of PeerJ, but as stated earlier this is not a literature review – instead it is an somewhat out of date opinion paper. The literature has not been reviewed rigorously or thoroughly, many of the papers cited are the authors’ own. Several times quotes are used but not referenced, and whole bold statements of fact are not referenced (i.e., lines 367 that children looking at butterflies in drawers increases their connection with nature and appreciation of biodiversity – this may or may not be an urban myth).

The organisation is fine – the sections make sense, but the ms is very wordy and ideas repeat in different sections.
Validity of the findings: The manuscript is not really novel – and the discussion of the role of museums has moved o a long way since 2017. In Europe the DiSSCo consortium (not primarily for digitising specimens as the author state!) has been accepted into the EU’s Infrastructure Roadmap and numerous symposia and presentations have occurred that are relevant here. In the USA iDigBio is transitioning from NSF funding to a new sustainable model, with some difficulty it might be said. The SYNTHESYS project (referred to in the ms in several different ways) has entered into a new phase of funding with new goals and research aims.

I am not convinced this paper has a well-developed argument that is current in this fast-changing landscape.
Additional comments: The ms is the result of a meeting held in 2017, but much has occurred since then. It would be important to really cite more of the relevant literature, and also be up to date with developments in for example DiSSCo and SYNTHESYS in the Europe, and iDigBio in the USA. I would have appreciated a bit of analysis as well as to uses of collections – there is a lot of information on this around, and were the paper a real literature review, there would be merit in pulling this together. But as currently written this is more of an opinion piece (outdated as well) than a literature review, and has a distinct bias to northern Europe and the USA. The recent symposium volume on use of collections in the Royal Society Transactions B, for example, does a better job of citing more of the relevant literature. All these ideas are being discussed at many meetings and symposia, and I am not sure of the audience of this manuscript. Is it to admonish museums? or garner support for museums? An example of this is in lines 497-8 and lines 506-7 which contradict each other, and no examples of this kind of activity e.g., reaching out to artists) are provided. There are just too many statements of fact that are not referenced or adequately backed up.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: THE GLOBAL MUSEUM: NATURAL HISTORY COLLECTIONS AND THE FUTURE OF EVOLUTIONARY SCIENCE AND PUBLIC EDUCATION
Review round: 2
Reviewer: 1

Output:
Basic reporting: This manuscript has benefitted greatly from the review process.  I want to commend the authors for their detailed consideration of the suggestions made by the reviewers.  Having now read through the revised manuscript, while I may not agree with all aspects of the manuscript it makes a solid contribution to the literature on this subject.
Experimental design: Nothing to comment on
Validity of the findings: The manuscript is well written and the revised version corrects concerns raised by the reviewers.
Additional comments: The responses to the reviewers have been handled very well.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: COMPLETE PLASTID GENOME SEQUENCES OF TWO SPECIES OF THE NEOTROPICAL GENUS BRUNELLIA (BRUNELLIACEAE)
Review round: 1
Reviewer: 1

Output:
Basic reporting: No comment.
Experimental design: No comment.
Validity of the findings: No comments.
Additional comments: It is normal paper to present  the plastomes. It is first paper that present plastome of Brunelliaceae. The features of plastome of Brunelliaceae were also analyses. The phylogenetic position of Brunelliaceae were also inferred.
 It would be better to inlude some photos of two species in the paper to illustrate the morphological difference to these two species.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: COMPLETE PLASTID GENOME SEQUENCES OF TWO SPECIES OF THE NEOTROPICAL GENUS BRUNELLIA (BRUNELLIACEAE)
Review round: 1
Reviewer: 2

Output:
Basic reporting: A:
Page(P) Line(L) 111, two species names are not italicized

P10L210 genus name is not italicized

B: these two papers should be cited for ((C,M)O) topology
Soltis et al. 2007 A 567-taxon data set for angiosperms The challenges posed by Bayesian analyses of large data sets

Exploring the phylogeny of rosids with a five-locus supermatrix from GenBank
Miao Sun, Ryan A. Folk, Matthew A. Gitzendanner, Stephen A. Smith, Charlotte Germain-Aubrey, Robert P. Guralnick, Pamela S. Soltis, Douglas E. Soltis, Zhiduan Chen
bioRxiv 694950; doi: https://doi.org/10.1101/694950
Experimental design: A:
Based on the abstract, the motivation and research goal is not stated, even though I read the answers in the introduction part.

B:
Since the phylogenetic framework of the flowering plants has been established by plastid genome data, I and maybe some readers are really curious why authors put so much efforts in the plastid genome, and just for two species?

C:
There are only 11 sequences in INFA.fasta file, far below the average; Including this alignment in the supermatrix would introduce extra gaps,  could bias the model estimation and phylogenetic tree inference.
Validity of the findings: A:
I feel like this work is solid but lacking of novelty and similar phylogeny from plastid/cp gene/genomes have been widely reported.

B:
The authors reconstructed phylogeny using 75 plastid protein-coding regions for 41 selected superrosid species. How this gene and taxon sampling schema is better than studies from (Ruhfel et al., 2014; 2014; Sun et al., 2015; etc). If the COM clade is heavily sampled, for 41 species, that means the rest clade of superrosid are sparse, which also means biased sampling, isn't it? Additionally, for three genomics in plant (mitochondrial, plastid, and nuclear), only sampled plastid is also not sufficient to resolve the inter relationship of OMC clade.
Additional comments: This manuscript is general well written, data is solid and analyses are statistically sound; all done in a start genomics and phylogenomic manner, however, I could not find the whole research motivation and results are exciting and appealing. Moreover, I think the authors should also justify their gene and taxa sampling is not biased. The era of application of plastid genome has passed, the whole tree of life in plant has been well established plastid genome, now the community should move on march to the biparental, more informative nuclear genes. Eventually plastid genome only tells one side of evolutionary story.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: COMPLETE PLASTID GENOME SEQUENCES OF TWO SPECIES OF THE NEOTROPICAL GENUS BRUNELLIA (BRUNELLIACEAE)
Review round: 1
Reviewer: 3

Output:
Basic reporting: I almost never review a manuscript so ready for publication. It is is clearly written and well explained.
Experimental design: This work has been done in accordance with the highest standards and best practices.
Validity of the findings: I have no doubts about the validity of these findings.
Additional comments: I have only these minor suggestions:

(1) Throughout, these authors use the past tense to describe observations that are currently true (although discovered in the past during the study), e.g., “Plastid genome size of B. antioquensis and B. trianae was 157,685 bp and 157,775 bp, respectively.” Although this is not uncommon in scientific writing, I personally judge that the present tense is more appropriate (while retaining past tense where appropriate, as when referring to something being done for the study, of course). This is just my personal preference and the authors are free to decide whether that would improve what is generally excellent writing.

(2) Line 201: Is the word “Brunellia” extraneous here?

(3) Lines 224 and 225:  The text reads, “All COM sequences analyzed had the same gene order and three Locally Collinear Blocks were 225 identified in the LSC, IR, and SSC (Fig. S1).” It is not clear at this point what taxa share this “Locally Collinear Blocks,” so perhaps this should be preceded by the listing of the other COM species from the literature are included for this statement.

(4) Table 1: I suggest replacing “C/G” with “C+G” and “A/T” with “A+T.”


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: COMPLETE PLASTID GENOME SEQUENCES OF TWO SPECIES OF THE NEOTROPICAL GENUS BRUNELLIA (BRUNELLIACEAE)
Review round: 2
Reviewer: 1

Output:
Basic reporting: This revision is in good condition. Most of problem has been resolved. The language is professional.
Experimental design: Good. It falls within the PeerJ.
Validity of the findings: Good.
Additional comments: The manuscript is ready for acception.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: COMPLETE PLASTID GENOME SEQUENCES OF TWO SPECIES OF THE NEOTROPICAL GENUS BRUNELLIA (BRUNELLIACEAE)
Review round: 2
Reviewer: 2

Output:
Basic reporting: As stated in my first review
Experimental design: As stated in my first review
Validity of the findings: As stated in my first review
Additional comments: Update review comments based on authors' revision:

The authors did a great job in their revision. I'm also glad to see the authors conducted additional analyses by excluding INF A region, and compared with their original results, and confirmed the results are not impacted.

I'm basically fine with this ms as it is, except one case as detailed below:

P10, L335-342: The authors stated that "Thus, the position of the COM clade remains uncertain (APG 2016).

1. I thought the phylogentic placement of COM clade has been nailed:  COM clade is embedded in malvids (maybe not monophyletic). Please check out recent 1kp paper (One Thousand Plant Transcriptomes Initiative, 2019): COM clade is nest in malvids in their ASTRL species tree analysis (Please see summarized rosid tree in the attached pdf).

2. Therefore I feel that this conclusion is out of date, and after APG IV (2016), there are so many studies pop out using nuclear data (e.g., Zeng et al., 2017; Leebens-Mack et al., 2019; Smith et al., 2019).

3. The authors should be clear that the Fabid clade is monophyletic only based on plastid data. Because it's not monophyletic, as nuclear data suggests COM clade and Zygophyllales are nested in malvids, respectively (Please see summarized rosid tree in the attached pdf). Please see Leebens-Mack et al. (2019) and Smith et al. (2019)

4. Thus I suggest authors should clearly declare in their results, discussion and conlusion sections that statements in present study only represent plastid data only (one side story from plant plastid genome). I know this study is only based on plastid data discussing phylogenetic relationships among taxa in rosids (as the authors fairly explained in the rebuttal letter ). However, the authors should acknowledge the new findings from nuclear datasets regarding to their focal taxa. The phylogenetic relationships are hard to be resolved if we only relied on data/information from only one part of three plants genomes.

FYI:
One Thousand Plant Transcriptomes Initiative, 2019:
Leebens-Mack, J.H., Barker, M.S., Carpenter, E.J. et al. One thousand plant transcriptomes and the phylogenomics of green plants. Nature 574, 679–685 (2019) doi:10.1038/s41586-019-1693-2

Smith et al., 2019:
Stephen A Smith, Nathanael Walker-Hale, Joseph F Walker, Joseph W Brown, PHYLOGENETIC CONFLICTS, COMBINABILITY, AND DEEP PHYLOGENOMICS IN PLANTS, Systematic Biology, , syz078, https://doi.org/10.1093/sysbio/syz078

Zeng L, Zhang N, Zhang Q, Endress PK, Huang J, Ma H. 2017. Resolution of deep eudicot phylogeny and their temporal diversification using nuclear genes from transcriptomic and genomic datasets. New Phytologist 214: 1338–1354.

One small thing:

In "SUPLEMENTARY MATERIAL" file, the genus name "Brunellia" should be italic in the first title


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: LINC00844 PROMOTES PROLIFERATION AND MIGRATION OF HEPATOCELLULAR CARCINOMA BY REGULATING NDRG1 EXPRESSION
Review round: 1
Reviewer: 1

Output:
Basic reporting: 1. There is a need to improve English. Many grammatical and spelling mistakes can be found throughout the manuscript.
2. Authors should provide an index to all the abbreviations used or expand them accordingly.
4. In the introduction, line 55, the information about the LINC00844 is insufficient. Authors should expand on this and include relevant information to emphasize as to why studying  LINC00844 is important.
5. In lines 61-62, authors should cite the appropriate manuscript to support their statement and the current citation(#16) does not support it.
6. Typological Error in abstract (LINC008444).
7. Line 151, 'markedly' should be changed to 'significantly' as the difference in LINC00844 expression is statistically significant.
8. Fig2A and 5A, please mention the p-value in the figure.
9. In line 163, what is LINC01554 ? hope it is a typo.
Experimental design: The primary research aim is within the Aims and scope of the journal.
The research questions are well defined. The experiments are performed with appropriate controls. The experiments performed in tissue specimens were collected in their institute with patient consent and ethical approval.
Validity of the findings: Authors present an interesting story of how LINC00844 regulates hepatocellular carcinoma cell migration by regulating the expression of NDRG1 and the results are novel to the field of HCC.

The article can be improved if the authors can address the following points.
1. Authors describe the methodology to identify differentially regulate lncRNAs but do not show the results for this analysis. In Figure 1A, they only show the expression of top 5 lncRNAs. Authors must include a figure/supplementary table of all the differentially regulated lncRNAs and the order of their ranking.
2. Figures 2B and 5B, regression analysis of LINC00844 and NDRG1 expression in different pathological stages should be performed and presented.
3. Authors should provide sufficient evidence to support to rational of exploring the relationship of LINC00844 expression with various clinical and pathological features described in Fig 3. Authors should also explain the biological relevance of all the differences in LINC00844 expression observed.
4. Fig 2H and I should be included in Fig 4. It is unclear to me as to why they have been placed there.
Additional comments: No additional comments


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: LINC00844 PROMOTES PROLIFERATION AND MIGRATION OF HEPATOCELLULAR CARCINOMA BY REGULATING NDRG1 EXPRESSION
Review round: 1
Reviewer: 2

Output:
Basic reporting: 1.	The authors need to rewrite or extend some parts as it is the case of the introduction. We suggest some additional references that might help to provide a better overview of the field and highlight the pivotal role of NDRG1 in non-pathological and pathological conditions.

2.	We strongly advise the authors to consider a proof-reading service which will substantially improve the overall quality of the manuscript.

References
1.	Luo Q et al. FOXQ1/NDRG1 axis exacerbates hepatocellular carcinoma initiation via enhancing crosstalk between fibroblasts and tumor cells. Cancer Lett. 2018 Mar 28;417:21-34
2.	Cai K et al.Ndrg1 promotes adipocyte differentiation and sustains their function. Sci Rep. 2017 Aug 3;7(1):7191
3.	Lu WJ, NDRG1 promotes growth of hepatocellular carcinoma cells by directly interacting with GSK-3β and Nur77 to prevent β-catenin degradation. Oncotarget. 2015 Oct 6;6(30):29847-59
4.	Jung EU et al.Hypoxia and retinoic acid-inducible NDRG1 expression is responsible for doxorubicin and retinoic acid resistance in hepatocellular carcinoma cells. Cancer Lett. 2010 Dec 1;298(1):9-15
5.	Akiba et al. N-myc downstream regulated gene 1 (NDRG1)/Cap43 enhances portal vein invasion and intrahepatic metastasis in human hepatocellular carcinoma. Oncol Rep. 2008 Dec;20(6):1329-35.
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The authors reported a novel and important role of LINC00844 in the onset of liver cancer. The authors addressed the hypothesis that LINC00844 might affect the levels of NDRG1 from different approaches by providing in vitro data as well as interesting data from human samples. The study is conceptually well designed and it would be suitable for publication in PeerJ, However, there are just a few minor points that need to be addressed.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: LINC00844 PROMOTES PROLIFERATION AND MIGRATION OF HEPATOCELLULAR CARCINOMA BY REGULATING NDRG1 EXPRESSION
Review round: 2
Reviewer: 1

Output:
Basic reporting: The authors have addressed all the comments and have made satisfactory changes. The manuscript is now written in professional English. It is my opinion that the manuscript is now acceptable for the broad readership of Peer J.
Experimental design: No changes required.
Validity of the findings: have addressed all the comments. No further revision required.
Additional comments: No additional comments


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: LINC00844 PROMOTES PROLIFERATION AND MIGRATION OF HEPATOCELLULAR CARCINOMA BY REGULATING NDRG1 EXPRESSION
Review round: 2
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The authors have extended the introduction part and have made a clear improvement in the quality of the manuscript.
However, in the methods part some headlines are wrong, the protocol for plasmids and lentiviral packaging and cell transfection contains a protocol for RNA isolation so it is inverted and it must be change. In addition, there are still a few grammar mistakes, thus, we strongly advise the authors to make a second proofreading to make sure the spelling and the use of the verb tenses are correct.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: TEENAGE SLEEP AND TECHNOLOGY ENGAGEMENT ACROSS THE WEEK
Review round: 1
Reviewer: 1

Output:
Basic reporting: 1.	Overall, the authors do an excellent job of conveying complicated methods clearly and concisely, and interpreting their results in a manner that is appropriate to the method.
2.	Measurement -
a.	In general, I found myself looking for more details in the measurement section, to better understand the nature of the many measures used and the extent to which they were endorsed by the participants.  I would have appreciated perhaps a table with means/standard deviations or even just the average rates for the different types of measures included in the text body itself.
b.	Line 232: The authors state that, they did not include measures of sleep duration as this would require assumptions they found untenable. Given the overall focus of the paper on including all possible specifications, I would like more detail on why this was untenable.
c.	It is admirable that this study includes such a wide variety of individual and family covariates and potential confounding variables.  However, I would have appreciated a bit more information/justification on why these variables specifically were included, and more detail on what these variables measure and how.  This is especially true for: parent word activity score (what does this measure? Why might it be a confound?), mother ethnicity (why not child ethnicity? How is this coded?), child long term illness (why is this included but other variables relevant to child wellbeing are not?).
3.	A few typos and wording issues were noted.
a.	Lower case standardized bweekend  rather than β in the abstract.
b.	Introduction line 61: “This engagement 61 oftentimes fail to cease” (should read “fails”).
c.	I would recommend rewording this sentence for clarity (line 300): “Such decisions researchers have to make to obtain an answer to their research question, quickly branch out into a vast ‘garden of forking paths’ that can skew the final results reported in a final paper”
4.	Figure 1 is a straightforward portrayal of many results. I would recommend indicating in more detail which variables were gleaned from self-report vs daily diaries (perhaps using asterisks as in the other figure) to aid the reader in deciphering patterns.
Experimental design: 1.	I greatly appreciate the authors' commitment to open science and transparency: They've made available via the OSF all study data and materials. This will greatly aid others in potential replication or extension of this work.
2.	As the authors note, the use of daily time use diaries is a significant contribution of this study over most of the literature which uses retrospective self-reports.  However, in deciding to take the average levels of reports across all study days (albeit separated by weekend and weekday), there is something of a missed opportunity in that we cannot draw conclusions about same day digital engagement and that same night’s sleep.  This decision not to use within-person multilevel modeling (and instead take a between-persons specification curve approach) may be reasonable given the study aims, but the implications of this decision ought to at least be mentioned and discussed in terms of future directions for research.
Validity of the findings: 1. I found that the study hypotheses were tested rigorously and transparently, and that results and their interpretations were appropriate in light of the method.
Additional comments: This paper was a pleasure to read- I think SCA is a good fit for this study question, and I would imagine this work will move the field forward in thinking carefully about the ways in which technology may (or may not be) linked with sleep difficulties.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: TEENAGE SLEEP AND TECHNOLOGY ENGAGEMENT ACROSS THE WEEK
Review round: 1
Reviewer: 2

Output:
Basic reporting: Overall well-written, with a clearly communicated rationale for the current study and recommendations for next steps.

Raw data are available via the UK Data Service and the analysis code is available (OSF) and clearly annotated.

Cites relevant literature throughout. The introduction summarises existing work on specifically bedtime media use (lines 90-94), but the discussion then argues that this is a gap in available literature with the current study " providing some of the first insights into the use of digital technologies before bedtime" (lines 522-524; similar at lines 619-620). This could be reworded to frame the current added contribution (of time use data and SCA) within the context of previous work (which has explored bedtime use).
Experimental design: This study adds a valuable new measurement for digital engagement via time use diaries. However, measurement of sleep parameters remains problematic and the limitations of this should be made explicit. In particular, the annotated code indicates that sleep duration has been calculated by assigning precise bedtimes and rise times based on ordinal response categories (e.g. assigning a value of 12.30am to all answers of 'after midnight' and 10.30pm to all answers of '10-10.59pm'). The available survey response categories do not allow accurate calculation of sleep durations. It should be made clear in the manuscript exactly how this measure was derived from the available responses, especially highlighting the limitations of using this rough proxy for sleep duration. Similarly, the manuscript should highlight the limitations of using the last sleep onset of the day as 'bedtime' (which may not be appropriate for those with more fragmented sleep), and also assigning midnight to any sleep onset later than this.

More generally, additional details on measures would be useful: both sleep and digital engagement self-report measures would benefit from further clarity on how they were derived (e.g. the composite score for digital engagement), how they were treated (e.g. as ordinal or continuous, and whether they met fair assumptions for this).

Similarly, the authors should mention their approach to handling missing data (e.g. listwise deletion?) and whether survey weights were used in analysis. The authors have not claimed that these analyses remain representative of the UK adolescent population, but these points can still be noted for transparency.

I would be interested to know whether the subsample (one third) of participants who gave time use data differed from the whole sample, e.g. demographic profile, self-reported sleep/media use.

The authors have provided considerable details on their use of SCA. However this is still likely to be an unfamiliar method for most readers and would benefit from even clearer communication. Including 'worked examples' could be helpful to illustrate concepts to readers (e.g. null dataset creation, bootstrapping p-value calculations). Furthermore, a large section explaining the bootstrapping and significance testing (lines 371-390) does not include citations - was the current approach based on similar previous work, or theory or is it a novel approach?

I appreciate the authors' transparency in outlining a detailed account of their approach, sharing clearly annotated code and noting the unsuitability of pre-registration for this study. I think this transparency could be further enhanced by clarifying the process of choosing possible specifications. For example, bedtime digital engagement could equally have been specified as 1 hour or 2 hours before bed, and/or as the amount of time using media in these periods rather than participation (yes/no). The same applies to other possible covariates that could have been included (or the choice of covariate grouping). Addressing this point may simply require explicitly highlighting within the manuscript that whilst these 120 specifications offer added insight (versus just one specification), they still do not provide comprehensive coverage, and that many other plausible specifications exist.

Is linear regression appropriate for predicting the ordinal outcomes? Were there any sensitivity analyses or citations that could support the suitability of this approach?

Were the correlations Spearman or Pearson (line 399 and Figure 1)? Were all reported correlation coefficients statistically significant?
Validity of the findings: The authors rightly point out the limitations of the 'screen time' construct (e.g. lines 100-102). This could be emphasised more consistently into the discussion, noting that the current analyses use an aggregated measure of different media use. This aggregates potentially very stimulating, socially driven online interactions together with potentially very passive media consumption. This limitation could be highlighted in the discussion, with the potential for future work to explore the role of content and experience of digital engagement, rather than hours per day.

Similarly, it is important to highlight in the discussion that bedtime digital engagement has been operationalised as the presence/absence of engagement in the 30 minutes before bed, rather than the time spent on this before bed. Results may differ between bedtime media users who spend more/less time on this activity.

Comparing weekdays and weekends is a valuable contribution, given the strong influence of school timetables on adolescent sleep schedules, as discussed (e.g. lines 538-544).
Additional comments: Thank you for this well-written manuscript that presents a valuable new approach to this topic and clearly highlights the issue of measurement as this field moves forward! I have provided details of areas where I feel the manuscript could be further improved, largely around providing additional clarity on the methods used and what these can and cannot yet tell us.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: TEENAGE SLEEP AND TECHNOLOGY ENGAGEMENT ACROSS THE WEEK
Review round: 2
Reviewer: 1

Output:
Basic reporting: Thank you for this opportunity to review this revised manuscript.  Overall, I found that the authors were quite responsive to reviewers’ suggestions, and the manuscript is improved as a result.  In particular, I appreciate the added detail in the measures section, which greatly enhances the clarity of the methods.  Nonetheless, and I hope the authors will forgive me here, I still have a number of questions about how some variables were extracted/calculated, and worry that readers may likewise have a hard time following elements of the procedures and how the different measures were computed (and thus how to interpret the findings):
1.	I appreciate the clarification in the rebuttal letter on my point 8 (about a missed opportunity to look at within-person effects).  I see that I mistakenly assumed that there were multiple time use diaries filled out on weekdays and weekends (which would have allowed for examination of within-person effects).  However, upon further review I realize that nowhere in the procedure is this important detail mentioned -that adolescents filled out only two-time use diaries (one on a weekday and one on a weekend).  I suspect that this missing detail contributed to my own misunderstanding and should be added to the procedures to avoid confusion on the reader’s part.  In fact, the procedure for the time use diaries in general could use a bit more detail: How/when did adolescents fill out the time use diaries? Did they always fill them out at a set time of day (e.g. 9pm), or rather at bedtime?  This also raises another question- given that the authors chose to use listwise deletion, I would be curious to know what percentage of youth who participated in the time-use diary sub-study contributed both weekday and weekend observations?  What was the final sample size before/after listwise deletion?
2.	The calculation of total time spent sleeping (from the retrospective self-report) remains unclear.  The revised manuscript details “To obtain the total time sleeping we first calculated the time spent sleeping before midnight: adding 7.5 to the adolescent’s score on the retrospective self-report bedtime question. We then calculated the time spent sleeping after midnight: for weekdays we added 4.5 to the score on the self-report wake up time question and for weekends we added 6.5 to the score on the self-report wake up time question. We then summed the time spent sleeping before and after midnight for both weekend and weekdays separately and used the measure as continuous. For the measure of total time spent sleeping on weekdays the mean was 8.62 (sd = 1.04) and on weekends it was 10.53 (sd = 1.23).”  Given that self-reported bedtime was reported using a likert scale (1-5), and not exact clock times, the procedure used for calculating the time spent sleeping before/after midnight is not straightforward.  The code (and Reviewer 2’s comments) suggest that discrete clock times were assigned (e.g. 10:30pm for answers of 10-10:59pm)- this detail ought to be included in the manuscript text itself rather than relegate to the code (which many readers will not reference). Furthermore, the decision to add 7.5, 4.5, and 6.5 needs more explanation.
3.	Time-use diary reports of sleep- the manuscript states that “We also measured adolescents’ bedtime using a measurement extracted from their time-use diaries, where one code signified the activity of sleep”.  What was the nature of the “activity of sleep”? The wording now sounds as if an adolescent reports on what time they fall asleep on Monday before they fall asleep- I am having a hard time wrapping my head around how an adolescent reports on what time they fell asleep on Monday (but does not do the reporting once they wake up on Tuesday- how does one report what time they fall asleep before doing so? Or maybe they do fill out the time use diary on Tuesday, about the entire preceding day?). The inclusion of the variable wording might make this clearer, as would more details about the diary procedure (see point 1 above). Also, the revised manuscript notes that the authors “imputed the time of last sleep onset during each diary day”.  The use of imputation here needs more detail.  What was the procedure for imputation?
4.	Retrospective self-report of digital engagement: the measures currently include the item wording as, “On a normal week during term time how many hours do you spend…”  Is this question meant to report on digital engagement over the course of the whole week?  Or rather on a typical day?  Furthermore, this section states that, “To obtain a composite digital screen engagement items we converted the measures to a 10 point scale, and then took the means, deleting those adolescents who had missing values in one or more of the four items.” How/why was the 1-8 likert scale converted to a 10-point scale?
5.	Time-use diaries of digital engagement: the measure section currently states that, “for those adolescents who did participate in digital screen engagement, we measured the total time spent on such activities”. For clarification, were those who did not report any digital engagement that day, were they coded as 0, or excluded from analyses using this “time spent” variable?   If they were not included, how did this affect the sample size given the use of listwise deletion?
Experimental design: No further comment
Validity of the findings: No further comment
Additional comments: No additional comments


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: METABOLIC RESPONSE OF SCAPHARCA SUBCRENATA TO HEAT STRESS USING GC/MS-BASED METABOLOMICS
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript is clear and well written in English with some minor grammar errors. However, the literature references are not sufficient which need to be developed more (please see details in the general comments). The discussion is not well organised.
Experimental design: The research question is clear and well defined and the experiment is well designed. However, there are some unclear points about the methods. It’s not clear about sample collection and preparation for metabolomics and there is no description of quality control of metabolomics analysis.
Validity of the findings: The study aimed to use GC-MS metabolomics to reveal insights into the impacts of heat stress on the clam S. subcrenata. The Data are well presented and results are novel and interesting that contribute to the understanding of temperature stress in marine invertebrates.
Additional comments: Line 64-65, please separate references for metabolomics and transcriptomics.
Line 74-77, it’s too general, could you please go more specific? Which are these environmental stressors? Pathogen infections, water contaminants, temperature? There are many GC-MS investigations for marine bivalves which have been published recently. A recent review paper by Nguyen and Alfaro (https://doi.org/10.1016/j.aquaculture.2019.734488) may help you to have an overview of the topic. In addition, a discussion on metabolomics studies on heat/temperature on marine mollucs/bivalves needs to be added. For example, a study by Alfaro and Nguyen on surf clam (https://doi.org/10.1016/j.aquaculture.2018.08.065). In general, this section needs to be expanded.
Is Young et al 2015 correct reference here for environmental stressor? Please check.
Line 77-79, I don’t think its correct for all environmental stressors. Again, you need to expand your literature review to see which environmental stressors have been studied with metabolomics approaches. Then, made a specific statement for the stressor and the organism that you are studying.
Line 92, please delete spectroscopy.
L109, how did you control the temperature when exchange water?
L111-114, should be mentioned in the introduction
L118, assay? It should be analysis or measurement. Please edit.
L133-134 do not match with lines 114-118. Please edit. Where did you get 30 mg weight samples?
L187, the mortality of what?
L132 and 147, Have did you do quality control for your analysis? Please see https://doi.org/10.1007/s11306-019-1556-8  as an example.
L188, 75.00%? please edit to make it consistent for the whole manuscript.
L217, “quantified” is this study used a quantitative method? How did you measure the concentration? I don’t see it in your method. Please check and edit.
L217-219, please reword, add “e.g.,”  after (, such as…. including amino acids (e.g., aspartic acid,…),…
Discussion: No clear separation between paragraphs, so it a little difficult to follow.
L240-243, please check grammar.
L244-245, which studies? Need references here.
L289, Is it a new paragraph? Where does it end? Could you please re-organize this paragraph with a clear statement of your findings following relevant references to support your findings?
L280, please also re-consider to add https://doi.org/10.1039/c8mt00092a which is relevant to this one.
L295, it more relevant to cite references of bivalve studies that found an increase of lactic acid and TCA cycle under stress conditions such as:
https://doi.org/10.1007/s11306-019-1556-8
https://doi.org/10.1016/j.aquaculture.2018.08.065
https://doi.org/10.1016/j.jip.2018.08.008
 In addition, more discussion about TCA cycle would be interesting since there are many recent studies in bivalves found the increases of TCA cycle intermediates.
Overall, based on your conclusions I suggest considering the following outlines for your discussion which a clear statement in each paragraph with supporting references.
•	energy metabolism,
•	osmotic regulation,
•	amino acid metabolism,
•	carbohydrate metabolism and lipid metabolism
Conclusion: May consider to add suggestions for future research?
Figure 5. What is the unit?


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: METABOLIC RESPONSE OF SCAPHARCA SUBCRENATA TO HEAT STRESS USING GC/MS-BASED METABOLOMICS
Review round: 1
Reviewer: 2

Output:
Basic reporting: The manuscript is written clearly with professional English used throughout, with the submission being ‘self-contained’ and representing an appropriate ‘unit of publication’.

The content of the introduction sufficiently and concisely describes the field background to demonstrate how the work fits into the broader field of knowledge and contains relevant sources of cited literature.

Overall the article is well-structured including figures and tables. Figures are relevant to the content of the article, of sufficient resolution, and appropriately described and labelled.

Raw data has been provided. I assume the metabolomics data provided has been pre-processed first (i.e., normalised to total peak area, internal standard, and biomass etc)? A clearer description of the GC-MS data provided would be useful.
Experimental design: The research is original primary research which fits within the aims and scope of PeerJ.

The research question is currently a bit ambiguous, although ‘omics studies do tend to be discovery-driven without prior hypotheses. This is OK in my opinion since the authors do state how their research fills an identified knowledge gap which is good.

The investigation appears to have been conducted rigorously and to a high technical standard – although further details on the quality of metabolite identifications are required to evaluate this properly. Please provide more information on how the metabolites were matched against the Fiehn database and what specific criteria were used. See my General Comments below.

Methods are mostly described with sufficient detail & information to replicate, but there are some areas which require more detail (e.g., metabolite identification process, statistical analyses criteria and reporting). See my General Comments below.
Validity of the findings: All underlying data have been provided; they appear to be robust (assuming the metabolite identifications are of high quality/standard).

However, the validity of the data and findings are currently difficult to evaluate, since the statistical methods need to be described in more detail and some justifications should be provided for particular method choices.See my General Comments below.
Additional comments: In this study, the authors provide a metabolomics-based analysis of clam mantle tissue after different levels of thermal stress treatment. The study provides new physiological information on how clams are able to adapt to thermal stress and could be a valuable contribution. While the manuscript is very well-written, there are a number of areas which require attention and clarification before a proper evaluation on the validity of the results and interpretation can be made. Most of these are centred around choices made for data processing and statistical analyses. Some specific comments follow, which could help to improve the manuscript.

Lines 133-134: Was the data normalised to sample-specific biomass at some point (or was each sample exactly 30mg)? This should be clarified.

Lines 133-134: Were the samples weighed frozen and wet? With biomass being based on wet weight? Please clarify.

Line 168: The authors normalize their data to the total peak area of the chromatograms. Authors should be aware that this technique assumes equivalent total metabolite signal per sample and is suitable only when the majority of all analytes remain constant. However, this ideal situation often does not hold due to the nature of the samples, therefore such normalization may distort data potentially masking true biological trends. It is more common to normalise the data to the internal standard, to account for systematic errors during metabolite extractions/derivatizations (e.g., pipetting variability, sample loss), potential derivative instability, and within- and between-batch variations (and preferably also using pooled QC samples). Please provide justification for the normalization approach used.

Lines 182-183: The selection criteria for metabolites were based on OPLS-DA VIPs >1.0 and t-test p-values <0.05. However, there is no mention of employing a method to control for multiple hypothesis testing during the univariate analyses. It is advised that this be controlled for using an appropriate technique (e.g., Bonferroni correction, False Discovery Rate threshold).

Line 161: Please provide more information on how the metabolites were matched against the Fiehn database and what specific criteria were used to do this. It is also advised that that the ‘Metabolite Identification Level’ be reported in the Methods section according to the recommendations of the ‘Metabolomics Standards Initiative’ (http://cosmos-fp7.eu/msi.html). This could be a good place to also state the ‘Metabolite Level’ certainty for the identifications.

Lines 168-179: Although the authors do perform proper model validations and report the relevant model statistics for their supervised multivariate analyses techniques, it appears they have not transformed and/or scaled their data prior to running these models. This is crucial for PCA/PLS-DA/OPLS-DA so that the data has normal probability density function, and all metabolites are given similar weighting during the analyses (see Berg et al. 2006 [DOI:10.1186/1471-2164-7-142]). Please provide justification for not transforming and/or scaling the data prior to PCA/PLS-DA and OPLS-DA.

Line 188: Survival was optimal at the highest temperature stress, and lowest at ambient temperature. These results seem counter intuitive. Some elaboration on these results would be useful in the discussion. Or has the list of survival rates been listed in the wrong order?... according to the discussion, this might be the case! Please double check this.

Lines 191-202: It is not clear which statistical analysis was conducted on these data. ANOVAs? Please clarify where appropriate.

Line 205: Here there is mention of QC samples. However, no mention of these were provided in the Methods section. Please include in the methods what these QC samples were and how they were prepared. Also, how were they used exactly for quality control purposes?

Methods Section: There is currently no information in the methods section for the pathway analysis that was performed. Please provide these details, and include which criteria were used (i.e., test statistic and measure of centrality and network topology, and which model reference pathway library was selected as a proxy for the marine mollusc focus in this study?). Which metabolites were entered into the quantitative enrichment analysis – all of them, or only the statistically significant ones? Also, the authors refer to the topology Impact Factor score as if has substantial relevance to the results, which it doesn’t – the authors should be interpreting these results based on the pathway analysis p-values in the first instance. The impact score is an indicator of how important the metabolites identified are to the network function only (is usually not changed by a stress treatment unless metabolites appear or disappear in one group or the other). A very high impact score can be obtained but if the p-value is also high then there is no treatment effect on the pathway. This should be reflected in the results and discussion when interpreting the outcome of this analysis. Note that metabolite transformation and/or scaling is usually performed prior to Pathway Analysis also.

Lines 229-235: This section requires some attention to better describe the results of the Pathway Analysis, with specific mention of pathways which were identified as being ‘statistically significant’.

Table 1: It is unclear how many components were used to obtain the R2(cum) and Q2(cum) values for these models. Please clarify. It is also not clear what the non-cumulative R2 and Q2 values represent for the OPLS-DA models – is this based on the permutation testing? Please clarify in the table caption.

Table 2: Only one pathway (histidine metabolism) shows signs that it may be affected (p<0.05) by thermal stress – but the impact score is zero which means that the metabolites identified are limitedly of importance to the pathway function. Consider this during interpretations. Note that it is good/transparent to also present the False Discovery Rate values for these pathways also, and the number of hits (i.e., x metabolites identified in the study which matched that particular KEGG reference pathway).

Table 3: No pathways appear to be significantly enriched according to their test statistics. Consider this during interpretations

Supplementary Tables 1 & 2: Please double check the fold change values. For example, 3-indoleacetonitrile in ST1 is currently listed as having a foldchange of 0.00. There are a number of similar entries in ST2 with highly significant p-values (P<0.0001) but foldchanges of zero which doesn’t seem right.

Discussion & Conclusion sections: These sections are generally well-written, and links the metabolite data with other works on marine invertebrate physiology. Although it seems that the pathway analysis does not provide much supporting evidence that numerous pathways were being affected, as stated in the conclusion

Other: I urge the authors to re-visit the pathway analysis components of this study. The authors have identified a very comprehensive list of metabolites, and I would have thought that many of the pathway impact values would typically have been higher because of this (?). Some suggestions in this area would be to 1) include all metabolites (significant and non-significant) in the analysis using their KEGG code IDs (can be time-consuming to obtain, but will best-ensure that good coverage across numerous pathways is obtained); 2) be sure to use an invertebrate pathway library (unfortunately there are no molluscan models and the researchers will need to select the most closely related taxa – perhaps C. elegans). Note that MetaboAnalyst v4.0 has recently updated their KEGG libraries for the Pathway Analysis modules.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: METABOLIC RESPONSE OF SCAPHARCA SUBCRENATA TO HEAT STRESS USING GC/MS-BASED METABOLOMICS
Review round: 2
Reviewer: 1

Output:
Basic reporting: The manuscript has significantly improved. The text is generally well written. I recommend it for publication.
Experimental design: Authors have added more details into the method section which made the manuscript is sufficient in details.
Validity of the findings: Data are well presented and results are novel and interesting that contribute to the understanding the effects of temperature stress on marine invertebrates.
Additional comments: Thanks for editing the manuscript according to my suggestions.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: LIGHT-HARVESTING CHLOROPHYLL A/B-BINDING PROTEIN-CODING GENES IN JATROPHA AND THE COMPARISON WITH CASTOR, CASSAVA AND ARABIDOPSIS
Review round: 1
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The results presented are mainly descriptive, and the results are lack of further evidence to be supported.

Although sufficient data were provided, the major problem is the following.
(1)	The results are relatively simple, and the main figures didn’t provide enough information. The authors identified genes by BLAST searching. Then the authors performed hierarchical cluster based on FPKM values. I feel the work didn’t use very strong evidence to tell what the expression pattern and molecular biology function of the genes. I suggest the authors test expression levels of genes in figure 4, and 5 using qRT-PCR.
(2)	The authors performed a synteny analysis in the text. However, no images in the main text related were shown.

Some minor concerns are as the following.
(1)	Log2 based values are observed in line 121, while log10 scale are observed in other parts.
(2)	Line 183. Arabidopsis should be italic.
(3)	Line 250. Was should be were.
(4)	Line 256. light harvesting should be light-harvesting.
(5)	Line 280. Subfamiles should be subfamilies.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: LIGHT-HARVESTING CHLOROPHYLL A/B-BINDING PROTEIN-CODING GENES IN JATROPHA AND THE COMPARISON WITH CASTOR, CASSAVA AND ARABIDOPSIS
Review round: 1
Reviewer: 2

Output:
Basic reporting: No comment.
Experimental design: The submitted article is within the scope of PeerJ. The study was well-conducted, and the results, especially for the gene identification and annotation, are quite solid.
However, many methods used are not fully described, in favor of pointing out to other articles. This is the case for instance on lines 96, 105, 107, 113, 116-117, 118, 122. The authors should at least reduce the number of these references and give a more thorough description of their methods.
Validity of the findings: No comment.
Additional comments: I only have a few other minor comments:
1.	Information about the functions associated to the conserved motifs can be found in the lines 191-200; this important information should also be added to Figure 3, where the motifs are depicted without further information.

2.	Concerning the expression profiles of the genes: in lines 215-219, the authors state that the genes can be divided into 5 clusters, depending on their tissue-specific expression pattern. A reminder of the genes, or at least gene families, present in each cluster would be welcome.
Also, do these clusters correspond to functional division? For instance, Cluster I, with a strong expression in leafage, seems to contain mainly stress genes, while the main light harvesting antennas of PSI and PSII are divided between Clusters II and V.
Developments of this part, in the results and/or discussion, would thus be welcomed.

3.	In the discussion, the lines 305 to 319 are just an overview of the role of some genes in stress response in general. A few lines on the role these genes might play in the particular stresses studied here (salt and drought) would be welcome.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: LIGHT-HARVESTING CHLOROPHYLL A/B-BINDING PROTEIN-CODING GENES IN JATROPHA AND THE COMPARISON WITH CASTOR, CASSAVA AND ARABIDOPSIS
Review round: 2
Reviewer: 1

Output:
Basic reporting: No comment.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: The manuscript is improved.
However I still have two minor concerns:

1. I appreciate the effort of the authors into supplying more info on the motifs in figure 3. However I believe that the functional information provided in the text in lines 210-220 in the track-changed manuscript (attribution of the motifs to CB domain, transit peptide, Rieske-like domain…), even though not available for all motifs, would be better suited and more understandable to non-specialist readers than just representative sequence in that figure.

2. Previously my last comment was the following: “In the discussion, the lines 305 to 319 [now lines 352-368 in the track-changed manuscript] are just an overview of the role of some genes in stress response in general. A few lines on the role these genes might play in the particular stresses studied here (salt and drought) would be welcome.”
Although the authors accept this comment and write that the manuscript has been revised, I cannot find this revision in the text: the section is identical to the previous version, and the only change to the discussion concerns the cluster analysis and makes no mention of salt and drought stresses.

I am satisfied with the other answers and revisions.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: LIGHT-HARVESTING CHLOROPHYLL A/B-BINDING PROTEIN-CODING GENES IN JATROPHA AND THE COMPARISON WITH CASTOR, CASSAVA AND ARABIDOPSIS
Review round: 3
Reviewer: 1

Output:
Basic reporting: No comment.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: I am satisfied with the modifications of the manuscript, and have no further comments.