The world’s oldest DNA to date: with the codes to a two-million-year-old ecosystem

By
Henrik Larsen
Kap København

A Danish-led, international research team has found and identified numerous ultra-short fragments of DNA in two-million-year-old layers of sediment deposit in northern Greenland.

This DNA is the oldest ever found and analysed, beating the previous record for prehistoric DNA – from a bone from a Siberian mammoth – by about a million years.

The ancient DNA was found on the surface of microscopic grains of mainly quartz and clay in 41 drill-core samples taken from the Kap København Formation: a deposit almost 100m thick near Greenland’s northernmost point.

MastalakeFN
Illustration of the landscape as it was 2 mio. years ago. By Beth Zaiken. 

The two-million-year-old DNA dates from a period when Greenland lived up to its name: when it was green and forested. The material collected includes DNA from plants, trees, animals and microorganisms, enabling researchers for the very first time to portray a prehistoric ecosystem based on DNA.

The climate in Greenland at that time varied between Arctic and temperate, around 10–17oC warmer than the North Greenland of today. This two-million-year-old material also gives us a picture of an earlier stage of development of the DNA of a range of species we are familiar with today.

Kap København i dag
Illustartion of the landscape as it looks today. By Beth Zaiken.

 

‘We can see the DNA of animals, plants and microorganisms, such as reindeer, hares, lemmings, geese, birch, poplar and thuya, from this early stage of development. There’s also DNA from species now extinct, such as the mastodon. And that’s pretty amazing because we’ve never previously been able to place the mastodon, a gigantic ice-age mammal, in Greenland. From the sea, we’ve found DNA from the Atlantic horseshoe crab and from green algae, both of which live in relatively warm marine waters. Of course, it’s highly fascinating to find and be able to study the details of such ancient DNA. But what’s equally incredible is that we’re suddenly able to peek directly into the engine room – into the very DNA – of a past ecosystem,’ say Professor Eske Willerslev and Professor Kurt H. Kjær, from the Lundbeck Foundation GeoGenetics Centre at the University of Copenhagen (UCPH).

The two professors, both specialists in ancient DNA, headed the study. It was recently published in Nature, one of the world’s most influential scientific journals, and the results will have implications for our understanding of evolution and climate change, among other things.

As the article in Nature concludes, the basic features of North Greenland’s prevailing climate two million years ago – around the transition between the Pliocene and the Pleistocene geological epochs – appear ‘to be similar to those of the climate expected to result from today’s global warming.’

En 2 million år gammel gren af lærketræ
A 2 million year old branch of larch tree still stuck in the permafrost within the coastal deposits. The tree was carried to the sea by the rivers that eroded the forested landscape.  Photo: Svend Funder.

A hump-backed deposit

But what did the landscape around Kap København look like two million years ago?

It was a low-lying coastal landscape, and its vegetation was, in many ways, similar to the terrain we see today at the northern Siberian tree line.

There was also an abundance of insects at Greenland’s northern point at the time. We know this due to earlier findings of insect remains in the Kap København layers and the fact that dryas, blueberries, field horsetail and a range of grasses were found among the 102 botanical genera whose DNA was identified in the 41 core samples.

The Kap København Formation was originally formed over a period of about 20,000 years from a mix of sediments from land and sea which accumulated at the mouth of Independence Fjord in the Arctic Ocean.

There was originally a shallow bay here, and the sediments at Kap København built up, metre by metre, into a hump-backed deposit during the course of the 20,000 years. DNA traces of the animals, trees, plants and microorganisms now described in the Nature article were contained in the sediment that caused the deposit to grow.

Ny optøet mos fra de permafrosne kystnære aflejringer
Newly thawed moss from the permafrost coastal deposits. The moss originates from erosion of the river that cut through the landscape at Kap København some two million years ago. Photo: Nicolaj K. Larsen.

And the genetic material has lain there well-protected ever since because, to date, the original layers of the deposit have mostly been covered by ice or permafrost. What’s more, these sediment layers have never been subject to human interference.

A total of 40 researchers from Denmark and a number of other countries, including the UK, France, Sweden, Norway, the USA and Germany, brought their specialist knowledge to the job of making the microscopic grains from the Kap København Formation talk in the Nature article.

Kjær and Willerslev explain that the process has been arduous and lengthy.

‘It all began in 2006 when the two of us worked together on a scientific expedition in Greenland. Expeditions are expensive, and you need to make the best possible use of your time and resources. So, when we were given the opportunity to pass by Kap København, we jumped at the chance. We collected a number of core samples from the large sediment deposit, and we’ve stored them ever since in the freezers at the University of Copenhagen. And it’s this material – plus some additional samples gathered in 2012 and 2016 – that forms the basis for the study and the article in Nature.’

From 2006 until pay-off finally came in 2021, the researchers at the Lundbeck Foundation GeoGenetics Centre made many attempts to examine the core samples, and they experimented with numerous different test set-ups and analysis techniques. But nothing helped because the grains of quartz and clay stubbornly refused to give up their secrets.

Not until a brand-new generation of equipment for DNA extraction and genetic sequencing hit the market a few years ago did the UCPH researchers – cautiously – begin to believe that the Kap København samples could actually have a unique story to tell. This equipment, which is constantly being refined, now enables researchers to locate and subsequently identify extremely small and severely damaged fragments of DNA, down to 30–50 base pairs (bp), in, for instance, sand and soil. So, these DNA fragments are merely a few millionths of a millimetre long.

 

In 2021, thanks to this equipment, and after a great many more tests, researchers achieved the final breakthrough in their search for Kap København’s original ecosystem. It was irrefutably clear: there were now scientific grounds for answering “yes” to two key questions they had been struggling with for years:

  • Is there any DNA at all in the two-million-year-old clay and quartz samples from the Kap København Formation?
  • And if they do actually contain DNA, can we detach it from the samples in order to examine it?

‘It was a pretty big moment when both questions were answered with a “yes”,’ say Willerslev and Kjær, ‘because we could now finally begin work to map a past ecosystem. It was a dream we’d had ever since we collected the first core samples from Kap København in 2006. But, to be honest, we’d doubted more than once along the way whether our dream would ever come true.’

 

Rester af trægrene fra skoven som voksede ved Kap København for to millioner år siden
Remains of tree branches from the forest that grew at Kap København some two millions of years ago. Photo: Svend Funder.

A helping hand to endangered species?

When you are standing looking “directly into the engine room” of North Greenland’s two-million-year-old ecosystem, it can be difficult – bordering on impossible – to decide which of the findings has the greatest significance. Detailed knowledge about the ancient ecosystem, with temperatures and species composition arising from a mix of Arctic and temperate climates, could indeed have implications for a number of fields. For example, Professors Willerslev and Kjær believe it would make sense to consider whether ancient DNA could contain valuable information about ecosystems’ reactions to climate change.

‘This is because the Kap København ecosystem, which has no present-day equivalent, existed at considerably higher temperatures than we have today – and because, on the face of it, the climate seems to have been similar to the climate we expect on our planet in the future due to global warming. One of the key factors here is to what degree species will be able to adapt to the change in conditions arising from a significant increase in temperature. The data from Kap København seem to suggest that, when subject to evolution, many species may be more “plastic” than we previously thought. That is to say, they may be tougher survivors thanks to their ability to adapt. And this is where the microorganisms come in,’ the two professors stress.

While reviewing the ancient DNA from the Kap København Formation, the researchers also found DNA from a wide range of microorganisms, including bacteria and fungi, and they are now in the process of mapping these. This work will be published in an upcoming research article, which will present a more detailed description of how the interaction – between animals, plants and single-cell organisms – within the former ecosystem at Greenland’s northernmost point worked, biologically speaking.

 

Eske Willerslev og Kurt H. Kjær er ved at blotlægge lagene
Eske Willerslev and Kurt H. Kjær are exposing the layers from the coastal deposits in connection with sampling for environmental DNA. Photo: Svend Funder.

‘In time, we may be able to use this knowledge to combat the depletion of biodiversity we’re currently facing,’ Kjær and Willerslev believe.

‘Biodiversity is already under intense pressure in many places due to global warming, and extinction may be on the horizon for some species, such as certain plants and trees. So, the strategy developed by plants and trees to survive in a climate characterised by rising temperatures – as was the case two million years ago at Kap København – could perhaps be used to make some endangered species more resistant to heat. For example, we could use genetic engineering to copy some of the two-million-year-old “tricks” from the plant DNA into today’s plants. It may also make good sense to ensure a specific soil balance by adding microorganisms with properties equivalent to those present at Kap København at that time. And we should also be able to achieve this by means of genetic engineering, if we choose to follow that path.’

 

 


Kap København

The exploration of the Kap København Formation began in earnest in 1979. Danish geologist Svend Funder visited the location and found remains of insects, plants, tree roots and mussels in the ancient layers.

Funder, who was attached to the University of Copenhagen and still conducts research there today, as an emeritus professor, has studied Kap København for more than 40 years.

The scientific community has long been aware of Kap København, and it was visited at the beginning of the 1920s by Danish geologist and polar explorer Lauge Koch.

However, very few other geologists have been there between the visits of Koch and Funder, and Funder was the first researcher to truly understand what an impressive witness to the era the deposit actually is.

Nuværende landskab ved Kap København
Present landscape at Kap København. The many hills have been formed by erosive dowm cutting of rivers cutting towards the coast. Photo: Kurt H. Kjær.

Salmon sperm lit the way

To a large extent, the 41 core samples from Kap København consist of quartz, which is a mineral, and of clay. Both the quartz and clay elements of the samples look like grains when studied under the microscope.

The ancient DNA proved to be stuck to the surface of these grains in the form of tiny fragments of genetic material from animals, plants and microorganisms.

The fragments measured mere millionths of a millimetre – typically 30–50 base pairs (bp) – and, thus, far from represented the complete DNA of the animals, plants and microorganisms in question.

The researchers had assumed and hoped that the DNA would be found on the surface of the grains. However, it was quite another thing to prove it. Extensive detective work was required, and Associate Professor Karina K. Sand from the Lundbeck Foundation GeoGenetics Centre at UCPH had the job of heading the investigation.

Sand, one of the authors of the Nature article, specialises in the interaction between organic molecules – such as DNA – and mineral surfaces. This interaction is highly significant for many life forms, and to understand it you need to be able to manoeuvre within the complex scientific intersection between geology, biology and chemistry.

Sand performed numerous experiments, adsorbing present-day DNA from salmon sperm on mineral surfaces equivalent to the material collected from the 41 core samples from the Kap København Formation – in particular, quartz and clay.

The aim of the experiments was to demonstrate how this DNA interacted with the mineral surfaces. Would it stick? If so, could it be detached? And would it be possible to extrapolate the DNA code from the surface of the microscopic grains?

Sand used a special microscopy technique – atomic force microscopy, which can see structures 1000 times smaller than those that can be observed under an optical microscope – to explore the questions, and she managed to prove that all could be answered with a “yes”.

The researchers then began looking for two-million-year-old DNA in the 41 core samples using a special device for DNA extraction.

Once the ancient DNA fragments had been identified, a new phase began. The researchers programmed their genetic sequencing equipment to compare every single DNA fragment with extensive libraries of DNA collected from present-day animals, plants and microorganisms.

Once this extensive programme had run, a picture of Kap København’s past ecosystem began to emerge in the form of the DNA of trees, bushes, birds, animals and microorganisms.

Some of the DNA fragments were easy to classify as predecessors to present-day species, others could only be linked at genus level, and still others originate from species impossible to place in the DNA libraries of present-day animals, plants and microorganisms.

 

Stubborn clay

Nærbillede af organisk materiale i de kystnære aflejringer
Close-up of organic material in the coastal deposits. The organic layers show traces of the rich plant flora and insect fauna that lived 2 million years ago at Kap København in North Greenland. The layers also revealed evidence of large mammals such as mastodons and reindeer using environmental DNA. Photo: Kurt h. Kjær.

It was definitely easiest to extract DNA from the quartz grains in the Kap København samples.

The clay grains proved to be much more stubborn.

This means that we can expect to be able to collect much more information about life at Kap København two million years ago if we can develop a method to force clay to release DNA, say Professors Kjær and Willerslev.

‘If we can develop such a method, the perspectives could be great – reaching far beyond the Kap København Formation. In general, DNA survives best in cold, dry conditions such as those that prevailed during most of the period since the material was deposited at Kap København. But we can’t rule out the possibility that clay may also preserve DNA in warm, humid environments such as those found in many locations in Africa. And if we can begin to explore ancient DNA in clay grains from Africa, we may be able to gather novel information about the origin of many different species – perhaps even new knowledge about the first humans and their ancestors.’