# Your Frogs for Science



## MonarchzMan (Oct 23, 2006)

I guess I'll toss out a feeler to see what folks thought of this idea.

I'm going to be doing phylogenetics on dendrobatids (mostly tinctorius, although I have some projects on the family in the works), and the hobby is actually a very large source of variety in dendrobatids. I have often thought that that could be utilized for investigative studies.

Essentially, the idea would be to get DNA from frogs to use in phylogenetic analysis. Typically, if I am doing this with wild frogs, I would take a liver sample or a toe clip, two methods that keepers would likely rather not do to their frogs. That said, though, there is access to DNA that hobbyists have that I normally wouldn't encounter with wild frogs: SLS froglets.

These frogs would be doomed to euthanasia anyway and tossed out, so why not put them to use for science? I have a few ideas at the moment for what I could do with samples from the hobby, but ideas would likely come to light as different types of frogs came in.

We talk about interest in science and how our frogs can be used for that, and I feel that this is a real, tangible way that the hobby can contribute to science. If you have frogs in need of euthanasia (SLS or otherwise), rather than just tossing them out, why not preserve them for use for scientific studies? Right now, they would be stored in in a -80C freezer until put to use, but better there than rotting, right? But there are a number of species that are poorly sampled for a variety of reasons, but that the hobby has good numbers of (e.g., terribilis).

All that would be required would be euthanizing them in a way that didn't degrade the DNA, getting a tube of ethanol (I'd send it to you), and then mailing the frog in ethanol back my way. I guess I am curious at the moment if there would be any interest from folks in participating.


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## Pumilo (Sep 4, 2010)

If I were dealing with someone trustworthy, where I know that this is a real research project that could help the hobby or our frogs in the wild in any way, I would be glad to participate. I think that is a great idea to use SLS frogs, or stunted froglets.
Obviously, I would consider you one of those people, and would love to participate in any way I could.


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## stemcellular (Jun 26, 2008)

Absolutely JP. I always have excess offspring that I would donate, sls or otherwise.


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## Rusty_Shackleford (Sep 2, 2010)

JP I can't say I would euthanize a healthy froglet for this purpose but I'd be happy to donate any SLS or otherwise unhealthy froglet for your research. I have a few different morphs of tinctorious that might help you.


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## frogface (Feb 20, 2010)

Could the eggs be used for this?


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## MonarchzMan (Oct 23, 2006)

Rusty_Shackleford said:


> JP I can't say I would euthanize a healthy froglet for this purpose but I'd be happy to donate any SLS or otherwise unhealthy froglet for your research. I have a few different morphs of tinctorious that might help you.


I wouldn't ask folks to euthanize froglets or frogs in their own collection, only those that would be condemned to death anyway, such as SLS froglets.



frogface said:


> Could the eggs be used for this?


I kind of doubt it, at least early stage eggs where the embryo has not grown. I'm not sure though. Not actually sure anyone has tried extracting DNA from frog eggs. Obviously, it would likely only have DNA from the mother (at least, that is what would likely show up), but beyond that, I'm not sure, to be honest.


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## frogparty (Dec 27, 2007)

Count me in. Anything in the name of science


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## jeosbo01 (Mar 1, 2004)

I think I can contribute a few here and there.


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## thedude (Nov 28, 2007)

Count me in as well. I don't really have any tincs though. I do have several froglets preserved in everclear though.


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## Pumilo (Sep 4, 2010)

Oops, I missed the "mostly Tincs" part. If it extends into thumbs, please feel free to hit us up.

Concerning the eggs, what if it were an obviously forming egg, or even tadpole? It seems like when Pumilio really get rolling, there are a number of abandoned tads now and then.


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## Mer_ (Sep 11, 2011)

I'd be happy to help if any of my froglets have SLS


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## Sherman (Oct 29, 2008)

Sounds like a good idea. I would happily donate.

I wish I didn't have anything that fits your criteria, but I do from time to time.


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## MonarchzMan (Oct 23, 2006)

Pumilo said:


> Oops, I missed the "mostly Tincs" part. If it extends into thumbs, please feel free to hit us up.
> 
> Concerning the eggs, what if it were an obviously forming egg, or even tadpole? It seems like when Pumilio really get rolling, there are a number of abandoned tads now and then.


Oh, it doesn't have to be just tincs. I would take any dendrobatid. Most of my work is going to be with tincs but I don't limit my questions to just those. I have broad interests 

There shouldn't be any problem extracting DNA from a tadpole, and like I said, I don't think anyone has tried eggs (fertile or not, mom's DNA is there). I think the only "problem" would be slicing off a portion of the egg since I wouldn't want to try an extraction on the whole thing lest the first time fail.


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## IEatBugs (May 15, 2012)

I too would be happy to contribute where I can. I have a few morphs of tincs and a number of auratus.


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## thedude (Nov 28, 2007)

MonarchzMan said:


> Oh, it doesn't have to be just tincs. I would take any dendrobatid. Most of my work is going to be with tincs but I don't limit my questions to just those. I have broad interests


Good to hear, I have mostly Ranitomeya to send you  Does it work that they are preserved in everclear and not ethanol? It works the same way and doesn't sever the DNA strands the way formaldehyde does.

When would you be doing this?


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## Ed (Sep 19, 2004)

thedude said:


> Good to hear, I have mostly Ranitomeya to send you  Does it work that they are preserved in everclear and not ethanol? It works the same way and doesn't sever the DNA strands the way formaldehyde does.
> 
> When would you be doing this?


 
Everclear is ethanol... 

Ed


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## MonarchzMan (Oct 23, 2006)

I could probably start it up as soon as next week since there seems to be lots of interest.


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## thedude (Nov 28, 2007)

Ed said:


> Everclear is ethanol...
> 
> Ed


Haha, Didn't realize it was just ethanol. Figured there were other ingredients in it.


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## frogface (Feb 20, 2010)

Ed said:


> Everclear is ethanol...
> 
> Ed


My state has pulled selling 190 proof. The highest proof they sell is 151. Is that adequate?


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## chuckpowell (May 12, 2004)

I'd like a better idea of what you propose to do. DNA analysis has been run on numerous D. tinctorius morphs and most other species. So what's your thesis? Where are you trying to find out? What experience do you have doing for the work you proposed? Just curious. 

Best,

Chuck 



MonarchzMan said:


> I'm going to be doing phylogenetics on dendrobatids (mostly tinctorius, although I have some projects on the family in the works), and the hobby is actually a very large source of variety in dendrobatids. I have often thought that that could be utilized for investigative studies.
> 
> Essentially, the idea would be to get DNA from frogs to use in phylogenetic analysis. Typically, if I am doing this with wild frogs, I would take a liver sample or a toe clip, two methods that keepers would likely rather not do to their frogs. That said, though, there is access to DNA that hobbyists have that I normally wouldn't encounter with wild frogs: SLS froglets.
> 
> ...


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## thedude (Nov 28, 2007)

frogface said:


> My state has pulled selling 190 proof. The highest proof they sell is 151. Is that adequate?


Ya that works fine, that was what I was told to use.

I would also like to add that I've drank both and the state banning one of them is useless because you still don't remember much the next day  But that's a story for the lounge...


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## Rusty_Shackleford (Sep 2, 2010)

thedude said:


> Ya that works fine, that was what I was told to use.
> 
> I would also like to add that I've drank both and the state banning one of them is useless because you still don't remember much the next day  But that's a story for the lounge...


Oh that's a story I'd like to hear....One shot for Adam, one for the frogs. Two shots for Adam, none for the frogs. Three shots for Adam...and pass-out.


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## frogface (Feb 20, 2010)

thedude said:


> Ya that works fine, that was what I was told to use.
> 
> I would also like to add that I've drank both and the state banning one of them is useless because you still don't remember much the next day  But that's a story for the lounge...


Thanks!

The reason they gave for banning the 190 was to make drinking safer for students, lol.


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## MonarchzMan (Oct 23, 2006)

chuckpowell said:


> I'd like a better idea of what you propose to do. DNA analysis has been run on numerous D. tinctorius morphs and most other species. So what's your thesis? Where are you trying to find out? What experience do you have doing for the work you proposed? Just curious.
> 
> Best,
> 
> Chuck


Definitely fair questions to be asked. Indeed, the literature is full of data on tinctorius, imitator, and pumilio, and even some other species (but relative to those three, it is minimal). Additionally, data available for those tend to be a few well know genes (cytochrome B, for example). You would be surprised, actually, as to how little is available, even for those big species. But such genes are not of the best of use, depending on the question asked. Cytochrome B is a very slow evolving gene, which is not actually of great use for looking at recent complexes, such as polymorphism.

I am mostly interested in how polymorphism can arise in aposematic species (conventional wisdom would say everything should be the same to better communicate signals to predators). Finding species such as pumilio or tinctorius are paradoxes to such ideas. Indeed, many dendrobatids break the mold. The question is why. What the hobby can offer is access to all of these complexes. I can examine polymorphism in a number of species, and compared phylogeography between different species (possibly predict where such complexes originated), and try to find a commonality in why a number of species are polymorphic and why some are not. Perhaps there are similar geographic origins of species that promotes polymorphism?

Like I said, it depends on what I get as to what questions I can ask. Depending on the variety of samples I get, it may even be possible to do some genomics and try to identify genes that control color and pattern.

Admittedly, I am new to phylogenetics, but I will be working with my adviser, Brice Noonan, who is pretty much the authority on tinctorius phylogenetics. So while I am new, I will have considerable experience helping me out and likely guiding me along the way. 

I hope that answers your questions.


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## hypostatic (Apr 25, 2011)

chuckpowell said:


> I'd like a better idea of what you propose to do. DNA analysis has been run on numerous D. tinctorius morphs and most other species. So what's your thesis? Where are you trying to find out? What experience do you have doing for the work you proposed? Just curious.


I agree with the quote above. The OP mentions a desire to perform a phylogenetic study, but that's about all the information that is given. I really feel that if you are asking for tissue samples from fellow hobbyists, then you should be as transparent as possible with your research plans and goals. If you were writing for a grant to the NIH to do cancer research, you wouldn't just hand them a sheet of paper that says you want to cure cancer. You would write a proposal including an abstract of the research you want to conduct, previous data published on the subject (by you or others), the merit of your research, how you plan on actually conducting the research (methods), how you wish to spend grant funds, etc...
While I recognize that Dendroboard is no NIH and that you're not asking for money or anything, I think that you really should provide this type of information to the hobbyists that are "funding" your research.
Here is the first grant proposal template google found, just so you have an idea of what sort of information you should provide to the hobbyists that are "funding" your research: Grant Proposal Template

Also, on the merit of the research, hasn't this research been done before? I know I've read papers on the subject before, and a quick google search brings up a few papers on the topic:
http://www.mvences.de/p/p1/Vences_A19.pdf
http://core.ecu.edu/biol/summersk/summerwebpage/articles/Brown2006_D.uakarii.pdf
http://www.asihcopeiaonline.org/doi...1(2006)6[623:PVSROL]2.0.CO;2?journalCode=cope
PoisonFrogPhylogeny
http://www.ias.ac.in/jgenet/Vol90No1/31.pdf
Rapid color evolution in an aposematic species: a ... [Evolution. 2008] - PubMed - NCBI
Refugial isolation and secondary contact in the dye... [Mol Ecol. 2006] - PubMed - NCBI
Molecular systematics and phylogeography... [Mol Phylogenet Evol. 2003] - PubMed - NCBI

Another question (there are so many haha), is why can't eggs be used? Not enough DNA in one cell? Would it be feasible to do something like a cheek cotton swab equivalent for the frogs?

And speaking of funding, another glaring question to me is where is the monetary funding from this research coming from? Last time I checked, enzymes and reagents aren't cheap, let alone sequencing. What facilities are going to be used for this?

Lastly, I would just like to say that I am not trying to inhibit the research in any way, and that I think every type of research is valuable (see:Revealed: The secrets of belly button fluff - Telegraph)


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## hypostatic (Apr 25, 2011)

Damn, you addressed it before my slow typing could ask the questions haha



MonarchzMan said:


> I am mostly interested in how polymorphism can arise in aposematic species (conventional wisdom would say everything should be the same to better communicate signals to predators). Finding species such as pumilio or tinctorius are paradoxes to such ideas. Indeed, many dendrobatids break the mold. The question is why. What the hobby can offer is access to all of these complexes. I can examine polymorphism in a number of species, and compared phylogeography between different species (possibly predict where such complexes originated), and try to find a commonality in why a number of species are polymorphic and why some are not. Perhaps there are similar geographic origins of species that promotes polymorphism?


Many frogs in the hobby have locality data, so they would be very useful in this purpose I feel. But also, I think many more frogs do not have this sort of data, and several times the exact reliability of the morph/locality of a frog can be questionable...

You know I was also wondering If you wanted to do some sort of analysis of the actual genetics of the frogs in the hobby, for example looking at things like gene flow, inbreeding depression, and general genetic diversity when compared to wild populations. Can this sort of thing be published?



MonarchzMan said:


> Like I said, it depends on what I get as to what questions I can ask. Depending on the variety of samples I get, it may even be possible to do some genomics and try to identify genes that control color and pattern.


What are all the questions that you would _LIKE_ to ask? 



MonarchzMan said:


> Admittedly, I am new to phylogenetics, but I will be working with my adviser, Brice Noonan, who is pretty much the authority on tinctorius phylogenetics. So while I am new, I will have considerable experience helping me out and likely guiding me along the way.


Ok, this sufficiently answers my funding/facilities question haha. University of Mississippi, correct? Are you a grad student working on your thesis?


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## MonarchzMan (Oct 23, 2006)

hypostatic said:


> Many frogs in the hobby have locality data, so they would be very useful in this purpose I feel. But also, I think many more frogs do not have this sort of data, and several times the exact reliability of the morph/locality of a frog can be questionable...


This is a potential problem, and again, depending on what I would get tells me what I can ask. If I get mostly frogs with poor to no locality data, I can use those for genomics (since I am interested in how genotype translates to phenotype). But if I can get samples with locality data, I can look at origins.



> You know I was also wondering If you wanted to do some sort of analysis of the actual genetics of the frogs in the hobby, for example looking at things like gene flow, inbreeding depression, and general genetic diversity when compared to wild populations. Can this sort of thing be published?


I am sure it can be published, and it is something I have thought about doing, but the only issue with that regard is spinning it so that it is not solely hobby related, if that makes sense. Knowing, for example, how inbred darts frogs are can have minimal impact and interest. If I spin it, though, to show how species can be maintained with X amount of allele loss in the private sector, then the audience gets much larger. That sort of study would require quite a bit of detail on the donated frogs, though.



> What are all the questions that you would _LIKE_ to ask?


Like I said, my main interests are around the idea of polymorphism. As far as I know, dendrobatids are probably the most polymorphic family of vertebrates. And I want to know why, since they shouldn't be. Are we seeing speciation or what? And this is where some genomics could play well. It could be that the only different between a red pumilio and a blue pumilio is a single nucleotide change (as seen in the spectral bears of the northwest). If it is that simple, single nucleotide mutation could explain the phenomenon. If could be that the gene that controls for color is rapidly evolving and under minimal selection. Who knows? But it all is quite intriguing.



> Ok, this sufficiently answers my funding/facilities question haha. University of Mississippi, correct? Are you a grad student working on your thesis?


Dissertation, actually. My work, like I said, will center around tinctorius (both phylogeography and field experiments), but my adviser always encourages new ideas (which is why I have proposals to work in Australia and Panama in addition to French Guiana and Suriname). I think that a broad sampling from polymorphic species will help tease apart the why. Get enough independent samples and some commonality will be made apparent (that is the hope, anyway).


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## easternversant (Sep 4, 2012)

Interesting project. I'd be interested in seeing how this turns out. Imitator doesn't seem to have much genetic sorting, but maybe tincs will.


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## oddlot (Jun 28, 2010)

Are you going to post your findings with us? Although I have only had 1 froglet with sls in the last few of years,I would participate if I produced any more.


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## MonarchzMan (Oct 23, 2006)

Yep, I'd definitely post results as I found them. I do want to make it clear, though, that it would take time since getting the required sampling for any given species would take a bit of time.


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