Proposal to evaluate genetic tools to evaluate Cattleya species

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Geoff, if the total DNA content by flow cytometry is going to be detectably different between species I can see that a tetraploid will have 2x the DNA of an established type species and be easy to identify and probably impossible to confuse with hybridization with any other species because none should have enough DNA to get that high. However, as we get to triploids and "mixaploids", I am thinking that we couldn't reliably distinguish these from hybrids with another species? They would be identified as "non-diploid" but we couldn't be sure what.
Terry, I agree that this would (hopefully) be most useful for telling if a plant is diploid or tretaploid and is likely the target species or not. There is some potential for people to have plants that are triploids (as a result of breeding 2n x 4n plants). This case would potentially require further evaluation of what the possibilities are for it being 3n or a hybrid. Could easily figure this out for plants with completely unknown parents which might include a root tip squash for confirmation. Mixaploids I think are pretty uncommon based on the data I've seen to date. This could occur at a higher rate in some of the really weird plants and explain variable behavior from bloom to bloom as the plant could have an unstable genome or actually be a chimera, but these would be very rare plants.

I agree the proposal has its limits in terms of what this information may or may not be able to do. My goal would be to make it easy for people to figure out if their plant is diploid or tetrapoid and likely the species or a hybrid (or something weird and unexpected). Certainly it will be imperfect in certain cases and may not be useful at all in others.
 
Got it. Establish the 2N DNA content with clear cut examples of a species that are not thought to be tetraploid or hybrids. Test some examples of the species that are thought to be tetraploid or possibly accidental or surreptitious hybrids.
 
Got it. Establish the 2N DNA content with clear cut examples of a species that are not thought to be tetraploid or hybrids. Test some examples of the species that are thought to be tetraploid or possibly accidental or surreptitious hybrids.
Spot on. I'm super curious what we will find. I'm also speaking with my friend at Illumina. They are the biggest gorilla in next generation sequencing. There may be additional data we could capture in parallel depending on the companies that do plant genetics/genomics since we will be gathering the necessary and interesting plant source material.
 
I’m interested to see what these labs can offer and what you intend to establish as the baseline plants of each species for comparisons.

Of course, you have my collection at your disposal. As well as my contacts in Japan, Asia and South America if need be.
 
As compared to chromosome counting, that requires cells in active division, I am thinking that we will want mature, resting cells with their diploid genome. We probably don’t want actively growing root tips or we risk too many cells that might have 4N DNA right before they divide. Do we just cut a piece of leaf and put it in a particular solution in a bottle and send it in?

To Leslie’s point, my statistics training pushes me to try and get about 11 scattered examples of the target species we pick. We want these to be as verified traditional/standard as possible, including even provenance that takes them to natural jungle collected. Natural collected doesn’t exclude tetraploids, etc, but it helps if everything else about the plant isn’t “excessive”. With 11, sort of randomly collected plants, we have a first shot at establishing a mid-point and range of the DNA content. Then we can compare with variant plants.

I am good friends with Jerry and Jason Fischer at Orchids Limited and I am sure that they would supply material from their substantial species collection.
 
I'll work on the details of what the ideal sample will be. You are correct Terry. We will likely need a small piece of mature leaf that isn't actively growing to minimize capturing a lot of cells with duplicating chromosomes. Orchids grow slowly, so even in such a case it probably would change things much. The idea of tracking down high probability diploid, true species plants in sufficient numbers is a very good idea. I think there are a number of people that we could enlist in this. Jerry&Jason would certainly have useful plants. Chadwicks and Fred Clarke come to mind as well.
 
Don’t forget David Off, Steven Kristoffersen, Lin Burzell, Jeff Bradley, Robert Fuchs and Ben Oliveros. Also Sergio Garcia and Michael Sinn, all with known jungle plants pre CITES.
 
I think Geoff and Leslie are best positioned to decide on a first test species. I think one with easy access to clear-cut diploid plants AND one that also has at least a little group of questionable plants that could be compared.
 
Only just saw this and read it with interest. Where did this end up?

Tim
 
Understanding which genome variations are responsible for adaptive phenotypes has been my line of work for the last 15 years. With collaborators from around the world we have been specifically interested in mapping colour pattern genes of poison dart frogs, and while we made important advances in pinpointing candidate genes…. the work is still ongoing. The reality is that bringing answers to these questions on frogs’ evolution has already cost a few millions USD, not even taking into accounts the salaries of senior investigators in France, USA, Germany, UK, etc.

What I want to point out is that while it is not impossible to identify the genes/mutations/pathways responsible for flower shape and colour variations, the scientific and monetary involvement is unfortunately out of the reach of our communities.

Sorry
Math
 
Understanding which genome variations are responsible for adaptive phenotypes has been my line of work for the last 15 years. With collaborators from around the world we have been specifically interested in mapping colour pattern genes of poison dart frogs, and while we made important advances in pinpointing candidate genes…. the work is still ongoing. The reality is that bringing answers to these questions on frogs’ evolution has already cost a few millions USD, not even taking into accounts the salaries of senior investigators in France, USA, Germany, UK, etc.

What I want to point out is that while it is not impossible to identify the genes/mutations/pathways responsible for flower shape and colour variations, the scientific and monetary involvement is unfortunately out of the reach of our communities.

Sorry
Math
That’s what I was afraid of.
 
Just to update as requested. Since i made this post, I have started two biotech companies and these have eaten up a lot of time.

That being said, I have downloaded most of the publicly available sequence data for Cattleya species genes and compared general themes that can be derived from the data. I've also looked at genome size info and have come to the conclusion that this might be useful for figuring out tetraploids cheaper than chromosome counts, it probably doesn't have enough fidelity to identify tweener plants - think walkeriana with loddigesii introgression or mossiae with lueddemanniana and certainly wouldn't be high enough to tell the difference between extremely closely related species.

From the data, it is clear that the unifoliate Cattleyas have distinct groupings and that a number of the species are of hybrid origin with genetic influence from one or more adjacent species.

My own groupings based on my interpretation of the genetic data:

1) Trianae series - quadricolor (candida), trianae, schroedae - These come from non overlapping habitats in three areas of Colombia. They are distinct from one another but effectively could reasonably be regarded as a species complex. Growth habit, rest and bloom periods are very similar with very large petals, often times overlapping. Over time have probably had contiguous and then separated habitat as global temperatures have oscillated over evolutionary time.

2) Lawrenceana series (lawrenceana, lueddemanniana, eldorado (wallisii) - These are distinctly different genetically from the rest of the unifoliate cattleyas suggesting that the center of evolution for this group was the isolated Guiana Highlands that then spread to the amazon (eldorado) and coastal Venezuela (lueddemanniana). These have distinct growth habits that are different from the rest of the unifoliate cattleyas and are generally smaller in size than other unifoliates. I tend to believe that lueddemanniana is essentially of hybrid origin from lawrenceana like and mossiae like progenitors. There is certainly ongoing gene flow from mossiae to lueddemanniana and vice versa today.

3) Labiata series - labiata, warneri, jenmanii - These are genetically quite similar to one another. Each has clearly evolved in non overlapping habitats that probably haven't ever been continuous. Each has distinct growth, rest and bloom periods that have been adapted to the local conditions of these distant habitats. Percivalliana and gaskelliana can also plausibly be added to this group although they likely have more influence from adjacent species than these three which are pretty well operated from one another and from most other species.

4) Luteola series - luteola, iricolor, mooreana - These are clearly closely related based on sequence data, but a bit uncertain of how luteola and iricolor came to be. Possible that they have slowly diverged with one another in a split at some point or they may have been moderately distantly related at one point and then have traded genes over time to become more closely related. Mooreana is absolutely of hybrid origin from these two species. Each of these is from the western part of South America in adjacent to overlapping habitats. All have relatively short lived, yellow flowers and powerful perfumes to attract pollinators. Luteola comes from the western Amazon basin at relatively low altitudes while mooreana and iricolor are more up in the mountains.

I tend to believe that the rest of the species are likely of hybrid origin and represent species with significant ongoing gene flow from adjacent species.

1) dowiana/warscewiczii group - There is obvious gene flow between dowiana/aurea with warscewiczii in northern Colombia. Some of the most beautiful and unusual cultivars of warscewiczii likely are from this introgression of genes. Dowiana/aurea on the other hand likely represent what happened when a purple flowered mountain cattleya migrated south into overlapping or at least nearby habitat to the plants that make up the luteola series. There is compelling genetic data that dowiana/aurea is a result of stealing some of the genes from progenitor plants of luteola. This is further supported by the fact that dowiana/aurea is generally warmer growing like luteola, has a powerful perfume and relatively short lived flowers. Rex likely represents the extreme of this as the furthest south down to Bolivia and has the least amount of influence from warscewiczii such that it has somewhat smaller flowers.

2) mendelli likely is the result of percivalliana and warscewiczii like plants creating a new species. I wonder if some of the turn of the century descriptions of mendelli having huge flowers if the result of a more introgressed portion of the population with warscewiczii. They were never very common. I wonder if the unnatural selection of these plants by collectors means that these genes are largely gone from the gene pool today and most mendelli clones today are somewhat more like the rest of the mountain cattleyas in size of plant and flower.

3) mossiae - this is clearly the result of the mixing of trianae, labiata and lueddemanniana like genes. It has features of all three both in growth and flowering as well as the fact that it is a highly variable species. The genetics supports this likely origin and the ongoing influence from lueddemanniana and percivalliana today. One could imagine that this represents the most distant colonization of the trianae group with signifiant introgression from the two adjacent species. Interestingly, both it and mendelli have the most similar growth pattern with flowering after a very long winter rest and then resuming growth in the mid to late spring. That probably has less to do with peculiarities of the habitat and more to do with some element of share origin.

As you can imagine, based on this assessment of likely how each of these species evolved and their somewhat overlapping genetic histories, it may be very challenging to come up with simple genetic markers to say if a plant is x gravesiana (mossiae x lueddemanniana) vs. an extreme of mossiae that has strong influence from lueddemanniana genes within mossiae that has been line bred to bring out those differences.

Based on this, I don't think my simple genome size approach is going to deconvolute very much and the work to sequence multiple genes and prove varying polymorphisms between the above groupings is also not very practical from a hobby standpoint to tell the difference between true species and hybrids. This is completely valid for larger population studies, but I agree is beyond the scope of what we could crowd source as hobbyists at this point.

I've looked at the bifoliates and they have a similarly complicated set of relationships of distinct species complex like groups and species of likely hybrid origin between more divergent species.
 
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