DNA from museum exhibits
An effective method of DNA analysis of animals preserved in formalin has been developed
Oleg Lischuk, N+1
American scientists have developed a method for sequencing historical biological samples preserved in alcohol or formalin. It allows genomic analysis of rare and extinct animal species. The results of the work are published in the journal Molecular Ecology Resources (Phylogenomics using formalin-fixed and 100+ year old intractable natural history specimens).
DNA analysis provides detailed information about the evolution of species, their taxonomic affiliation, as well as the appearance and loss of various traits. However, the only samples of many rare and extinct species available for study are exhibits from museum collections preserved in alcohol, formalin or a mixture thereof. This method of preservation allows you to preserve samples for centuries, but greatly complicates the extraction of genetic material for sequencing due to chemical modification of tissues and DNA fragmentation. Therefore, until now, genomic analysis of such samples has been laborious and ineffective.
To improve the method of DNA extraction, the staff of the University of Louisiana selected samples of 21 rare snakes (17 different species) collected from the 1870s to 1990. 16 of them were stored in formalin, the rest in alcohol. A fragment of liver tissue weighing from 100 to 200 micrograms was taken from each of the canned snakes, without causing significant damage to the exhibits.
The tissue samples were kept for six hours in distilled water to remove the preservative residue. The crushed samples were then incubated for 15 minutes at 98 degrees Celsius with a standard detergent-based ATL buffer causing tissue lysis. After cooling, the protein-destroying enzyme proteinase K was added to the solution several times and incubated for another 48 hours at 65 degrees Celsius (until no visible tissue fragments remained in the solution). After such preparation, DNA was extracted by the standard method, but by increasing the exposure time, lowering the temperature and reducing the amount of extraction buffer. This method made it possible to obtain DNA suitable for research from 10 samples, including more than 100 years old.
This DNA was analyzed by sequencing a new generation, for which the nucleic acid must be divided into relatively small sequences. Since the DNA of the samples was fragmented during the preservation process, no additional processing was required.
Genomic analysis made it possible to identify more than 2,300 ultraconservative elements – non-coding DNA sequences that remain absolutely unchanged during evolution and are completely identical in different, even distant species. They were compared with the results of genomic analyses of modern snakes, whose tissue samples were taken specifically for sequencing. This made it possible to determine the position of rare and extinct species included in the study in the phylogenetic tree of reptiles.
The position of the studied samples in the phylogenetic tree.
Figure from an article in Molecular Ecology Resources.
"The importance of this work is that it has made species that have been practically lost to science due to extinction, rarity or a secretive lifestyle available for study in the era of modern genomics," concluded one of the researchers, Sara Ruane.
"It also highlights the importance of museum collections [of preserved specimens] for modern science," added Christopher Austin, the head of the work.
Modern methods of DNA analysis allow us to use a variety of biological samples and find answers to questions that were previously considered unsolvable. As a small number of examples, we can cite the proof of the existence of Neanderthals 400 thousand years ago, the determination of the place of domestication of single–humped camels, the unraveling of the origin of the clothing and food of the "ice man" Etzi - such enumeration can be continued for a very long time, and the amount of information received is growing literally every day.
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26.01.2017