A Neanderthal bone, a mummified Egyptian cat, a dog from Siberian permafrost, a sturgeon from Roman times or medieval plant remains: their DNA is a fantastic source of information. But the genetic code of these ancient organisms is not easy to crack. New methods, collectively known as next-generation sequencing, make biological collections like the one of our Institute more and more relevant.
DNA barcoding researcher Gontran Sonet welcomes us in one of the three labs for DNA research. Our colleague is currently using DNA extraction methods on dog bones, to gain expertise on the new methods for ancient DNA analysis.
This is the world of DNA sequencing. In short, DNA sequencing is determining the order of the nucleotides – the building blocks A (adenine), C (cytosine), G (guanine) and T (thymine) – of a DNA molecule to obtain the genetic code of an animal or plant.
The most widely used method – until recently – to do so was developed by Frederick Sanger in the late 1970s. In his method, the nucleotides (building block of the DNA molecule) at all positions of a specific DNA fragment are determined in a dedicated reaction using enzymes and synthetic short stretches of DNA, which have been designed for one specific location of the genome.
The obtained genetic information can tell us a lot about the organism. Specimens from museum collections or straight from archaeological sites can in this sense be the missing piece to a puzzle of the past. We can discover the diet of historic human populations, map migration routes of early Europeans, where and when the dog or dromedary was domesticated, how a disease spread or why one species went extinct and others did not.
A Big Net
The big barrier in researching ancient DNA – also called aDNA – however, is that it is horribly difficult to obtain information from it. The quantity and quality have decreased with time: it became fragmented in increasingly smaller pieces, and it has been contaminated by its environment or by parasites. To put it in perspective: 1 gram of fresh organic material contains about 1 microgram of DNA, while 1 gram of an ancient specimen contains only 0,0001 to 0,0000001 micrograms of DNA.
When you try to sequence aDNA using the Sanger method, it yields poor results. But that is where next generation sequencing (NGS) steps in: with new, more advanced methods it has become possible to sequence enormous amounts of DNA strands at once in parallel. And it is even possible with very short fragments. The moment you find a minimum overlap of thirty or more base pairs, you can analyse and reconstruct the DNA molecules. Gontran calls NGS ‘the big net’. It is expensive, but you catch a lot of information in return.
To show where DNA is prepared for that big net, Gontran leads us to the top of our Institute. There, on the thirteenth floor, is the third DNA-lab. Contrary to the two others, which are spacious and where light falls through the big windows, this lab is dark and small: there is barely enough room for two people to sit. To protect the precious DNA, the researchers have taken plenty of precautions. The door is password-protected. Inside there is a separate room for the scientists to change clothes and put on masks and lab coats. In the lab itself, there is a special air circulation system and UV-light to destroy unwanted DNA.
Cat Mummies
Next generation sequencing has opened a whole new world for scientists: bones and fossils that were of little use to DNA-researchers in the past, are suddenly a goldmine of information. Claudio Ottoni from the University of Leuven, for example, uses DNA obtained from cat mummies to reconstruct the history of domesticated cats.
Ottoni is one of the 6 experts who presented at a workshop on NGS and ancient DNA, in our Institute last June. Members of a Belgian Network for DNA barcoding (BeBoL) and European experts shared their experiences on aDNA from archaeological bones, museum mammals, old herbaria and museum insects.
“We really have to follow a strict protocol. In some cases, only 0.1% of the DNA extracted from a tooth really belongs to the animal that we study”, says Katerina Guschanski, another expert at the workshop. The researcher from the Uppsala University in Sweden studies evolutionary relationships and speciation in guenons. Because it is hard to find the little monkeys, and because one third of the taxa are endangered, she had to find samples of specimens in museum collections, including our collection and the one of the Royal Museum for Central Africa. “Natural history collections are of immeasurable value in research like that”, she says.
The Future of Ancient DNA
Gontran expects that sooner or later, NGS will be confronted with its limits. “At the moment, NGS produces “big data”, allowing applications that were almost unconceivable previously. But we still need to standardize and cross check our procedures to make sure that results are reliable. Only when we have done that, we will know what is possible and what not.”
That is why he tempers the enthusiasm about some exciting scenarios regarding the future of aDNA research: the sequencing of very degraded DNA such as in dinosaur bones is not (yet) an option. “It seems that DNA has an expiration date. The DNA in dinosaur remains is at least 65 million years old, and at that age it has become so fragmented that it has become impossible to reconstruct the genome.” The ‘de-extinction’ of animals from the past, like a mammoth or a dodo, also remains fiction for now. “DNA is not everything: the animal that would be born out of an elephant with reconstructed mammoth DNA would be a genetically modified organism, still missing many biological characteristics of a mammoth.”
But if we look at the possibilities today, it becomes clear that our Institute plays an important role in modern DNA-research. As the curator of a collection containing more than 37 million specimens, the RBINS offers countless opportunities for new studies. “In JEMU – a research unit of the Royal Belgian Institute of Natural Sciences and the Royal Museum for Central Africa – we want to improve our skills in this field, because we work directly with museum collections”, Gontran says.
The Joint Experimental Molecular Unit (JEMU) is an integrated research infrastructure funded by the Belgian Science Policy and supported by the Royal Belgian Institute of Natural Sciences (RBINS, Brussels) and the Royal Museum for Central Africa (RMCA, Tervuren). JEMU aims at supporting scientific research on natural history collections in the fields of DNA barcoding, phylogeny reconstruction and archiving biological specimens. The JEMU team is composed of Massimiliano Virgilio, Carl Vangestel, Nathalie Smitz, Gontran Sonet and is coordinated by Thierry Backeljau and Marc De Meyer.
