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In their paper on the molecular genetics of the Sinningia alliance, Perret et al. used two data sets and then a combination of the analyses of those two. The general idea was to compare restricted pieces of the genomes of all the available species. One looks for DNA which has not been undergoing selection, either positive (increasing the frequency of beneficial mutations) or negative (decreasing the frequency of deleterious mutations). If the DNA segment is not an active part of any gene or regulatory region, it should be free to "drift", and the number of differences in the region between one species and another should be one measure of the genetic distance between the two.
Almost all the DNA in multicellular animals and plants is in chromosomes contained in a bag within the cell called the nucleus. These nuclear chromosomes get mixed-and-matched as part of sexual reproduction. For their analysis, Perret et al. used four introns from a single gene.
Chloroplasts, the cells that produce chlorophyll and do the photosynthesis that turns sunlight into energy for the plant, have their own DNA, separate from the nuclear DNA. Plastid DNA is not mixed during sexual reproduction, since pollen grains do not contain chloroplasts. Therefore plastid DNA is inherited only from the seed parent.
From the plastid DNA of the Sinningieae, Perret et al. used four spacer regions between genes and two introns, each one from a different gene.
The plastid DNA segments were quite a bit longer overall than the nuclear DNA segments. They also showed more differences between species. There were no differences in the selected nuclear DNA segments between some species pairs, such as S. piresiana and S. rupicola, which are almost certainly very closely related, and S. douglasii and S. cardinalis, which are almost certainly not each other's closest relative.
The following table shows how well analysis of each of the DNA types and a combined analysis did in pairing up some "natural" species couplets.
| Species pair | Plastid DNA | Nuclear DNA | Combined |
|---|---|---|---|
| S. conspicua and S. eumorpha | closest | close but not closest | closest |
| S. cardinalis and S. magnifica | closest | in a large group of similar species | very close (in a group with one other species, S. lateritia) |
| S. cardinalis and S. bulbosa | very close (in a group with two other species, S. lateritia and S. magnifica) | close (closer than cardinalis-to-magnifica) | very close (in a group with two other species, S. lateritia and S. magnifica) |
| S. pusilla and S. concinna | not close | very close (in a group with one other species, S. aghensis) | not very close |
| S. gigantifolia and S. cochlearis | closest | very close (in a group with one other species, Vanhouttea fruticulosa) | closest |
| S. schiffneri and S. gerdtiana | closest | closest | closest |
| S. guttata and S. lindleyi | far | closest | far |
| S. douglasii and S. nivalis | close but not closest | close but not closest | close but not closest |
| S. carangolensis and S. valsuganensis | closest | closest | closest |
It should be noted that "closest" doesn't necessarily mean close. It just means nothing else is closer. S. schiffneri and S. gerdtiana aren't particularly close (not nearly as close, for instance, as S. leucotricha is to S. insularis); they are just isolated, with no other near relatives.
(In fact, there is a phenomenon called long branch attraction in this sort of mathematical analysis that makes the placement of branches which show a lot of evolution less reliable. It is therefore possible that S. schiffneri and S. gerdtiana and Paliavana plumerioides are not close relatives, but only wound up together by being excluded from the other groups of closer relatives.)
It's interesting to speculate about why the plastid DNA gets pusilla/concinna and guttata/lindleyi "wrong". Since chloroplasts are inherited only from the seed parent, they are not necessarily representative of the plant's overall heredity, but just one particular chain of ancestors. In the case of these sinningia pairs, hybridization might be the source for one of the species in each pair. If S. concinna were descended from S. harleyi x pusilla, for instance, it could have harleyi-like plastid DNA but pusilla's dwarf characters.
Of course, the plastid DNA could be getting it right, and we are being too impressed by physical similarities.
Peter Shalit has written that it was very difficult to get S. pusilla and S. concinna to cross, and that the resulting hybrid was almost infertile. The fertile "micro" hybrids were probably the result of tetraploidy.
So it seems likely that these two species, while close, are not that close.
Mostly, genes contain the DNA code for making proteins. The building blocks of DNA are called base pairs; the building blocks of proteins are amino acids. Each group of three base pairs in the gene corresponds to an amino acid. The cellular machinery takes the DNA code and turns it into a protein.
You would think that the code for a single protein would have to be a continuous sequence, with no interruptions, but such is not the case. Most genes have gaps of "non-coding DNA". These gaps are called introns. Introns get copied just like the "coding DNA", but are sliced out later.
The opposite of introns are exons. These are the active parts of the gene; the cell's protein-making machinery slices out the introns and uses only the exons to do its work.