RECONSTRUCT PART OF THE GENOME
Here is my prediction: Within the next couple of years, George Church and the mammoth revivalists will succeed in transferring at least one mammoth gene into an elephant stem cell. They will use that stem cell to produce cells that express the newly inserted mammoth gene. They will carefully measure their success by designing a smart experiment to show that the gene is now producing mammoth proteins rather than elephant proteins. When they see positive results, indicating that they have, indeed, engineered a mammoth gene into an elephant cell, they will announce this success with deserved pride. It will be an astonishing achievement.
No elephants will have been harmed in the process. No elephants will have been involved in the process, other than to donate blood during a routine veterinary visit. No female elephants will have been subjected to any experimental manipulation whatsoever. No one will have performed nuclear transfer on an elephant. No baby elephant whose genome contains mammoth genes will be gestating anywhere.
The press will not hear any of the above caveats, however. The headlines will read: “The Mammoth Is Back.” “Extinction Is No Longer Forever.” “Scientists Create Woolly Mammoth in Test Tube.” It will be the biggest, most exciting, scariest, most wonderful, most terrible thing to happen in recent memory. There will likely be widespread announcements of dire consequences as well as excitement and some hysteria.
But there is no need, really, to speculate on how people will react. We can simply look to recent history.
THE MAMMONTELEPHASE OF DE-EXTINCTION
On April 23, 1984, the following appeared in the Chicago Tribune, tucked neatly away among the inside pages. The headline read: “A shaggy elephant story.” The full article is reproduced here, with permission:
When a species becomes extinct, we expect it to stay that way. Scientists in America and the Soviet Union have upset that seemingly safe assumption by “retro-breeding” a hybrid animal that is half elephant and half woolly mammoth, The story starts in Russia, where Dr. Sverbighooze Yasmilov of the University of Irkutsk was able to extract the nuclei from egg cells taken from a young mammoth that was found frozen in Siberia. Technology Review reports that he sent the material to the Massachusetts Institute of Technology, where Dr. James Creak mixed the DNA from the cells with elephant DNA. Woolly mammoths, which roamed Europe until they died out 10,000 years ago, have 56 chromosomes; elephants, their near-relations, have 58. Based on Creak’s success, Yasmilov decided to try to fuse the nuclei from the mammoth’s egg cells with sperm from an Asian elephant. The experiment produced eight fertilized eggs, which were implanted in Indian elephants. Six miscarried, but two hybrid animals—males that are probably sterile—were born. The hybrids, which some call “mammontelephases,” are covered with yellow-brown hair and have jaws that are similar to the mammoths.
The tiny story was picked up and distributed by the Chicago Tribune’s news service, and versions of it appeared in more than 350 newspapers within the following days. It even appeared in a nationally circulated Sunday supplement, where it no doubt received the widest potential readership.
Not one of the newspapers that picked up and ran the story bothered to check the facts. If anyone had bothered to contact the author of the report mentioned in the Technology Review, for example, or had tried to talk to any of the scientists involved in the research, they would have made a startling discovery: the whole thing was a joke. The scientists did not exist. The project did not exist. The story was meant to be a parody of science, written by a talented undergraduate student to fulfill a science writing assignment. The story was published in the Technology Review in celebration of All Fools’ Day. The article, which is on page 85 of the April 1984 edition of the Technology Review, concludes with the name of the student author—Diana ben-Aaron—and the date—April 1, 1984.
Perhaps those at the Tribune and the many other newspapers that decided to run the story were simply too excited about the possibility of mammoth de-extinction to notice the date or to question the authenticity of the report (including the unlikely collaboration between Soviet and American scientists at the peak of the Cold War). Or perhaps they didn’t get the joke.
The fictional piece by ben-Aaron was prescient in many ways. She predicted, for example, the poor success rate of nuclear transfer, despite writing the article more than twelve years before of the birth of Dolly at the Roslin Institute. She predicted that the Asian elephant would be used as a surrogate, although it would be more than two decades before we knew with certainty that the Asian elephant is more closely related to the mammoth than the African elephant is. She also anticipated and attempted to defuse some of what the public would fear about de-extinction. For example, she foresaw that containing these new creatures—not allowing them to escape and breed with the wild elephant population—would be a key concern. As Michael Crichton would do six years later, she invented a mechanism by which breeding of the cloned animals would not be possible without human intervention. While Crichton’s dinosaurs were all female and therefore unable to reproduce, ben-Aaron’s mammontele-phases were all infertile males. She made them infertile by giving them an uneven number of chromosomes. With mismatched numbers of chromosomes, they, like mules, would be sterile.1
The reaction to the press coverage of ben-Aaron’s fictional article was fast, heated, and mixed. Some people celebrated, either because they were amused by the display of obviously poor journalism, or by the parody itself, or because they didn’t know it was fake and simply were excited that a mammoth had been brought back from the dead. Others were angry, either because they felt that the parody was improper or unfair or because they didn’t know it was fake and were really annoyed that scientists would do such a terrible thing as bring a mammoth back from the dead.
The reaction was, in fact, much like the reaction I anticipate when the mammoth revivalists publish the first evidence that their genome-engineering project has been a success and that edited elephant cells may—at some point in the future—be used to make edited elephants. Of course, the 1984 scenario was entirely fabricated. The hypothetical headlines of our future will reflect actual science going on in an actual cutting-edge research lab at one of the most respected research institutions in the world.
In 1984, those who read and believed the story in the Chicago Tribune or elsewhere came away with one message: a mammoth had been brought back to life. That was, however, not what the article said.
The headlines to come when the mammoth revivalists produce the first mammoth-flavored elephant cell are likely to be more spectacular than the subdued title of the Tribune piece. Careful journalists are unlikely to omit the fact that very little of the elephant genome is actually changed; however, this fact will be conveniently brushed aside to make way for impassioned and melodramatic commentary reflecting on the central message of the piece: a mammoth will have been brought back to life.
Except it still won’t be true.
IF IT LOOKS LIKE A MAMMOTH AND ACTS LIKE A MAMMOTH, IS IT A MAMMOTH?
Let’s return to the work that is going on in the present day. It is feasible today to use genome-engineering technologies to directly edit DNA sequences within a living cell. George Church’s lab is using this technology to edit elephant cells so that the genome within them looks more mammoth-like than elephant-like. For now, this work is limited to editing only one or a few genes in somatic cells. However, we have somatic cells that contain genomes in which some genes have had the elephant version removed and replaced with the mammoth version. This is the status quo of mammoth de-extinction.
If the somatic cells edited by the mammoth revivalists are used to create a baby elephant, that baby elephant would have only a very tiny amount of mammoth DNA. The mammoth revivalists’ goal is to engineer an elephant so that it can survive better in the cold. Let’s imagine that they achieve this by replacing the elephant version of something in the range of five to ten genes with the mammoth version of those genes. In this scenario, the phenotype of the hypothetical baby elephant hopefully would change, but more than 99.99 percent of its DNA would still be elephant DNA.
In the fictional scenario published in 1984, the babies that were born were first-generation hybrids, created by fusion of DNA preserved in a mammoth egg and DNA from elephant sperm. The hybrid creature’s DNA was 50 percent elephant and 50 percent mammoth, but ben-Aaron never went so far as to call them mammoths. In fact, she provided an entirely new scientific name—Elaphas pseudotherias—which places the hybrid mammontelephase in the same genus as the Asian elephant, but gives it an entirely new, and fictional, species name. Perhaps her goal was to be scientifically precise about what she created. Perhaps it was to avoid confusion. Whatever her motivation, the piece provides an excellent opportunity to observe the public’s reaction to the creation of a (fabricated) hybrid species.
The public did not care that it was a hybrid. The media called it a mammoth, and so it was a mammoth. Perhaps what was most important was how it was described, but even this was absolutely minimal in the media reports: the hybrid had yellow-brown hair and mammoth-like jaws. Clearly, even a tiny bit mammoth-like was good enough for the public. It was a mammoth.
This is great news for those in favor of de-extinction, because it provides an enormous amount of wiggle room for determining when de-extinction is a success. A mammoth will not have to be pure in order to be received as a mammoth. This is a relief, because—as we’ve discussed—while 100 percent mammoth is out of the question, 1 percent mammoth may not be.
This provides an opportunity to redefine de-extinction, shifting away from a species-centric view. Genetically pure mammoths, or genetically pure versions of any extinct species, are likely not possible. However, we do not need genetic purity to benefit from de-extinction technology. If we select wisely which 1 percent of the genome to change, we may be able to resurrect those characteristics that distinguish a mammoth from an elephant. More importantly, we may be able to resurrect those characteristics that allow the elephant to live where the mammoth once lived. Once released into the wild, the hybrid elephant could stomp around, knocking down shrubs and consuming vast quantities of vegetation. It could disperse seeds and insects and distribute nutrients. This new hybrid animal could replicate the mammoth, without necessarily being a mammoth, with vast potential benefits to the arctic ecosystem.
Most people who are seriously considering either de-extinction or back-breeding are doing so because they believe that bringing these species back would provide an upper hand in present-day struggles to preserve biodiversity and maintain healthy ecosystems. Extinctions at any level—whether of prey species or predator species or species that distributed seeds or species that consumed shrubs and trees so as to preserve open spaces—can have cascading effects across an entire ecosystem.
The project to breed back the auroch in mainland Europe aims to create giant herbivores that will graze wild, open land and thereby keep the shrubs and trees at bay. The result, the team hopes, will be a restored habitat that can be used by both large and small mammals and at the same time increase the diversity of plant species on the landscape. The auroch is the target phenotype of their back-breeding experiments. However, the team’s intention is not to bring an auroch back to life but to resurrect a phenotype that can do in that environment what the auroch used to do. They hope to replace the auroch with something similar in function but not necessarily identical in form.
In my mind, it is this ecological resurrection, and not species resurrection, that is the real value of de-extinction. We should think of de-extinction not in terms of which life form we will bring back, but what ecological interactions we would like to see restored. We should ask what is missing from the existing ecosystem that could be recovered. De-extinction is perhaps better imagined as an elaborate bioengineering project in which the biological end product is modeled on something that evolution created but that has unfortunately been lost.
WHAT PARTS OF THE GENOME SHOULD WE ENGINEER?
Genome engineering, and not cloning by nuclear transfer or back-breeding, seems to be the most likely avenue to resurrect extinct traits and—depending on how loosely we care to define a species—extinct species. But where do we begin? The answer to this question is likely to be different for each de-extinction project.
If our goal is to create an elephant that is capable of surviving a Siberian winter, then we have to change this tropically adapted species into something that fares well in the bitter cold. Longer, thicker hair will definitely help, as will hemoglobin with higher efficiency in carrying oxygen at low temperatures. But what other traits should we try to engineer? Are there other ways to make an elephant more efficient at maintaining its internal body temperature? Are there energetic requirements to living in the Arctic that we haven’t yet considered? Are there adaptations to the digestive system that will be necessary to allow an elephant to eat the plants that grow in Siberia? Do we need to engineer morphological changes that make the elephant capable of digging plants out of the snow? Will we need to engineer the elephant’s immune system so that it can evade pathogens that are not present in the tropics? These are all good questions to which we do not yet have any answers, much less a target gene or suite of genes that we could sequence and look for mammoth-specific changes that we will want to engineer.
The scientific world is unlikely to prioritize elephant genomics in the near future, which means that we won’t know anytime soon where all the genes are in the elephant genome, what these genes do, or how they interact with each other. This information, however, is all crucial if we really want to genetically engineer a mammoth in a piecemeal fashion. Given that so much is unknown, one solution might be to change every nucleotide in the genome where the mammoth differs from the elephant. In doing so, we would be less apt to overlook any important difference or interaction between genes. This would, however, require making a lot of changes: if we assume that the Asian elephant and the mammoth diverged from their common ancestor around four million years ago and that the rate of divergence is similar to that in other mammals, we can expect something like 70 million genetic differences between the two species (on the same order as the number of genetic differences that separate humans and chimpanzees). Less than 2 percent of the elephant genome would need to be edited, but 70 million changes is a lot of changes to make.
So how would we make those changes? First, we have to figure out what they are. Many (if not most) of the differences between the Asian elephant genomes and mammoth genomes can probably be identified by sequencing and assembling both genomes, lining them up, and scanning them for sites at which they differ from each other. Since we know that we will not be able to sequence and assemble a complete mammoth genome, we’ve already stumbled upon the first problem with this approach. Ignoring that problem, the next step is to design a strategy to change each of the elephant sites that differ into the mammoth version using genome-editing tools. If we assume that each edit will require its own CRISPR-RNA (the CRISPR-RNA is the part of the CRISPR/Cas9 system that finds and then binds to the part of the genome where the edit is to be made), then we need to design and deliver into the cell 70 million different CRISPR-RNAs. However, George Church’s lab has been improving techniques to insert larger and larger fragments of DNA at once, which may allow us to change multiple bases at the same time. Let’s assume that the technology gets really good, and we can make, on average, ten changes with each CRISPR-RNA. This would reduce the number of CRISPR-RNAs necessary to around 7 million.
In their mammoth hemoglobin work, George Church’s revivalists designed two CRISPR-RNAs to make three changes to the hemoglobin gene (one CRISPR-RNA made a single edit and the other made two edits). Editing the elephant sequence takes place in three steps: First, everything necessary to edit the genome—the CRISPR-RNAs, Cas9 (the molecular scissors), and the mammoth template DNA—has to be delivered into the cell. Second, the CRISPR-RNAs have to find the part of the genome that they are intended to cut. Third, the cellular-repair machinery has to paste in the mammoth version of the gene.
Because the mammoth revivalists have actually performed this experiment, we can use their results to estimate the overall efficiency of the cut-and-paste process. In other words, we can ask, what proportion of edited elephant cells end up with all three changes? The mammoth revivalists found that each CRISPR-RNA had a different efficiency in targeting the right part of the genome (the “cut” step), and that the cellular machinery had a different efficiency in fixing each cut the way we want it to be fixed (the “paste” step). In this experiment, they estimated that the cut-and-paste efficiency of one of their CRISPR-RNAs was about 35 percent, and the other (the one that makes two changes) was about 23 percent. This means that only 8 percent of cells ended up with all three changes.
Even if we were able to reduce the number of CRISPR-RNAs that we need to make to, say, 100 (many fewer than the 7 or 70 million estimated above), and we assume, generously, that the efficiency of each of these is somewhere around 30 percent, that would mean that we would have to change at least 5 × 1053 cells in order to find one cell in which all 100 changes were made at the same time. That’s a big number. To put this in some perspective (although perspective is very hard at this scale), scientists have estimated that there are around 40 trillion (4 × 1013) cells in a human body and 7.5 × 1018 grains of sand on Earth.
Fortunately, we may be able to narrow down the number of changes we need to make without resorting to targeting specific traits. First, some of the species-specific differences that we observe when we compare one Asian elephant genome and one mammoth genome will not exist if we were to compare all mammoths and all elephants. These sites will look at first like they differ between species because we have only a single individual of each species to compare. But, if we were to have multiple genomes from each species, we would notice that some differences are not fixed within either species but are instead variable among mammoths or among elephants. Since not every mammoth or not every elephant has these changes, we can conclude that these changes are not important in making a mammoth look and act like a mammoth (or making an elephant look and act like an elephant). We could therefore exclude these sites from genome editing.
Another way to limit the number of necessary edits is to make only those changes that occur within genes. The genome is a big place, and only a small portion of the genome—around 1.5 percent of the human genome, for example—is made up of genes that code for proteins, while the rest of the genome is made up of other, noncoding DNA. Because genes code for proteins and proteins make phenotypes, the most important genetic differences between two species might be those that are found within the sequences of the genes themselves.
There are, unsurprisingly, several problems with this strategy. We do not, for example, know the location of all of the genes in the mammoth genome, so finding them will require educated guesswork—comparison with better-studied genomes—and even then we may not find all of the genes. Also, targeting only those differences that occur within genes may miss important differences that are found in the noncoding portion of the genome, such as differences that change when or how much of a gene is expressed. Differences in gene expression can result in different phenotypes even if the sequence of the gene itself is exactly the same.
Perhaps, then, we will need to make every change in the genome sequence. George Church believes that this will soon be feasible. The key, according to George, is to reduce the number of CRISPR-RNAs by cutting and pasting very long—very, very long—fragments of DNA. Instead of making a only few changes with each CRISPR-RNA, we will need to make thousands of changes, if not tens of thousands of changes, at once. Right now, George’s group is able to synthesize strands of DNA that are 50,000 base-pairs long. While the accuracy of such long synthetic sequences is still less than ideal, the technology is improving while the cost is going down. If it were possible to synthesize the entire mammoth genome in, say, 100,000 base-pair chunks, then we could cut-and-paste the entire mammoth genome into an Asian elephant genome using fewer than 350 CRISPR-RNAs.
Still, 350 is a big number and, following the logic above, would require an absurd number of cells even if each cut-and-paste experiment worked with exceptionally high efficiency. The logic presented above is not particularly logical, however, and does not describe how we would perform the experiment in reality. Rather than try to luck into a scenario in which 100 (or 350) things that are unlikely to happen all occur at the same time, we would perform the experiment in steps, where a few changes will be made and validated, and then a few more introduced to those cells that were edited successfully, and so on. The experiment would still be challenging, and it would still take a long time to complete, but it might just be feasible.
Today, we do not know the complete genome sequence of the mammoth. However, we are likely to learn most of the mammoth genome sequence within the next few years. Today, we cannot edit an Asian elephant genome so that it looks entirely like a mammoth genome. This technology is also improving. In fact, this particular step in the de-extinction process is probably the fastest moving in terms of technology development.
MORE THAN THE SUM OF ITS NUCLEOTIDES
Genome editing will become an increasingly efficient way to transform all or part of a genome of a living species into something that resembles the genome of a species that is extinct. However, some important differences between species may have nothing at all to do with the sequence of their genomes. Simply changing the genome sequence might not, therefore, be sufficient to resurrect the extinct phenotype.
Genomes are complicated places. Genomes live in cells, which live in bodies, which live in environments. And in different cells, different bodies, and different environments, the same genomes—genomes that are identical in both the coding and noncoding portions—can produce very different phenotypes. Identical twins, for example, have identical genomes. But, as they get older, identical twins become more and more phenotypically and behaviorally different from each other. How can this happen, if their genomes are the same?
In addition to their genome, all organisms have what is called an epigenome. The epigenome is a confusing concept, and not all scientists define or describe the epigenome in the same way. As I understand it, the epigenome can be thought of as a suite of tags that that are attached to the genome. These tags indicate whether a gene is turned on (making proteins) or off (not making proteins). Importantly, these tags are not actually part of the genome, which means they can be in flux throughout the organism’s life. Epigenetic tags can be heritable—that is, the epigenetic state of a particular gene is sometimes passed on from parent to offspring. These epigenetic tags might tell a cell to turn on only those genes necessary for being a heart cell, for example. Other epigenetic tags are not heritable in this traditional sense and, instead, may appear or change because of interactions that take place between the organism and the environment in which that organism lives.
A variety of environmental stimuli are known to affect the epigenome. An organism’s diet, the amount of stress or toxins it is exposed to, and how much physical exercise it gets will all alter the epigenome, changing which genes are expressed, when they are expressed, and how much they are expressed. By the time identical twins become adults, their epigenomes differ considerably, although their genomes remain identical. It is the combination of their genome sequence and the epigenetic differences that accumulate over each twin’s lifetime that results in their distinct phenotypes.
Will epigenetics complicate de-extinction efforts? We don’t know. If we edit an elephant gene to contain mammoth DNA sequences, it will, as it begins to develop, contain the elephant epigenome. In the womb, it will be exposed to the developmental environment of an elephant: a mom that eats an elephant diet, lives in the elephant habitat, and expresses elephant genes. It will survive by virtue of the elephant placenta, which will be expressing elephant genes modified by that particular mother elephant’s epigenome.
While we cannot study the effects of the developmental environment using identical twins (because they develop in the same intrauterine environment), we know the health and diet of the mother during pregnancy can have profound effects on fetal development. Her diet can even affect health outcomes later in life, such as risk of heart disease and obesity. Fascinatingly, we also know that the mother’s diet before pregnancy can influence the epigenetic state of her genes, with consequences to the developing embryo. Almost certainly, the diet and amount of stress to which the mother elephant is exposed will affect her developing mammoth (or mammoth-like) embryo, but exactly what these effects will be remains unknown.
In some instances, a species-specific developmental environment is not critical to a successful gestation. Robert Lanza’s genetic-engineering firm, Advanced Cell Technology, successfully cloned both a gaur and a banteng (both living but endangered species that are closely related to cattle) using nuclear transfer and female domestic cows as surrogate mothers. Both pregnancies went well, and both calves appeared to thrive. It is unclear, however, how these animals might differ from clones that were born from surrogate mothers of their own species.
What about the environment after birth? Epigenetic changes accumulate throughout life and are driven by the environment in which an organism lives. How much of looking and acting like a mammoth is due to having a mammoth genome, and how much of it is due to living life in the steppe tundra? This is something we may have to wait to learn.
Understanding the genome and how the genome interacts with the environment is among the major technical hurdles standing in the way of successful de-extinction. It is unclear today how this hurdle will be surmounted. Will we finish sequencing the mammoth genome and learn where all of the genes are and what all of the genes do, so that we can make a minimal number of changes and still end up with a mammoth? Or will genome-editing technology advance to the point where we can make all the changes necessary to create a genome that is 100 percent mammoth-like? Will we devise a way to infer the epigenetic state of ancient tissues, as a first approach to learning which genes should be turned on or off in unextinct individuals?
Answers to these questions may come soon. Knock-in and knock-out experiments—where scientists either turn on or turn off specific genes in organisms like yeast, fruit flies, and mice—are being used to discover where genes are, what they do, and how they interact with each other. Large, population-level human genome-sequencing projects are being used to identify specific genetic changes that are associated with distinct phenotypes, such as adaptation to life at high altitudes or susceptibility to cancers or other diseases. These experiments are homing in on ways to identify what are likely to be the most “important” changes to make. At the same time, the technology behind CRISPR/Cas9 systems is developing rapidly. These systems have so far been used to edit the genomes of more than twenty different species, chopping out and inserting fragments of the genome that are on the order of tens of thousands of nucleotides long. We may eventually arrive at a solution where it is possible to edit an entire genome.
Ancient epigenomes may even be within reach, thanks in part to how DNA degrades over time. It turns out that DNA methylation, which is one way that the epigenome marks the genome, interacts with DNA degradation in an interesting and useful way. In methylation, the epigenome modifies the genome by attaching a methyl group (CH3) to a cytosine—one of the four nucleotide bases that make up DNA. DNA degradation also affects cytosine bases, but in a different way. Cytosine bases are often deaminated as DNA degrades—they lose part of their chemical structure (an amine group) and become uracil, which is a nucleotide base that is otherwise not found in DNA. When methylated cytosine bases become deaminated, however, the interaction between the two chemical modifications converts the cytosine into thymine, another of the four nucleotides found in DNA, rather than uracil. The ancient epigenome can be reconstructed by distinguishing deaminated cytosine bases that become thymine bases (which degraded after being tagged by the epigenome) from those that become uracil bases (which also degraded but were not epigenetically tagged).
This approach was first used by Ludovic Orlando’s research group at the University of Copenhagen in Denmark to reconstruct the epigenome of a 4,000-year-old Paleo-Eskimo from Greenland’s Saqqaq culture. Soon afterward, a team of scientists from the Max Planck Institute for Evolutionary Anthropology in Leipzig, Germany, and the Hebrew University of Jerusalem, Israel, mapped the epigenome of two archaic hominins—a Neandertal and a Denisovan. The team found around 2,000 differences between the reconstructed epigenomes of the archaic hominins and the epigenomes of modern humans, some of which may underlie some of the skeletal differences between us and our archaic cousins.
While technologies to sequence, edit, and understand genomes are all developing at a rapid pace, new tools that become available tend to work best for those species that are the best studied. Far less is understood about elephants than about mice, fruit flies, or humans, and the same is true for many of the candidate species for de-extinction. These tools can be adapted for research on other species, but, for now, the hurdles standing in the way of fully reconstructing the genomes of extinct species remain high. George Church, however, is very tall.
Plate 1. Martha, the last known living passenger pigeon, in her enclosure at the Cincinnati Zoo in Ohio, USA. Photo courtesy of the Wisconsin Historical Society, WHI-25764.
Plate 2. Bones of mammoths (first), reindeer (second), bison (third) and horses (fourth) collected along the banks of the Kolyma River, Duvanniy Yar, Siberia. All of the approximately 1,000 bones depicted here were collected in a single day and over an area of about 1 hectare. Photo credit: Sergey Zimov.
Plate 3. Leg bones from three passenger pigeons whose genomes are being sequenced at the University of California, Santa Cruz, as part of the passenger pigeon de-extinction project. These were among the remains excavated by Dr. Greg Sohrweide from a site in Onondaga, New York, USA, and date to the 1690s. Photo credit: Andre Elias Rodrigues Soares.
Plate 4. Sorting the remains of stingless bees from fragments of ancient amber in the ancient DNA facility at the Pennsylvania State University. Although amber-preserved insects were once thought to harbor preserved ancient DNA, research has shown that DNA does not survive in amber, even over relatively short periods of time. Photo credit: Mathias Stiller.
Plate 5. Field sampling of ice age bones. Only a small amount of tissue is required for DNA extraction and analysis. Here, a small fragment of bone is removed from a sample collected on the Taimyr Peninsula, Siberia, during our 2008 field season. Photo credit: Beth Shapiro.
Plate 6. Placer mining near Dawson City, Yukon Territory, Canada. Here, gold miners blast the frozen soil with high-pressure water to expose the gold-bearing gravels beneath. As the soil is washed away, bones, teeth, tusks and other remains are revealed and can be collected. Photo credit: Tyler Kuhn and Mathias Stiller.
Plate 7. The partial skull of an ice age horse recovered from an active placer mine near Dawson City, Yukon Territory, Canada. Photo credit: Tyler Kuhn and Mathias Stiller.
Plate 8. A cervical vertebral bone from a mammoth is slowly exposed by placer-mining activities near Dawson City, Yukon Territory, Canada. Sometimes, several bones from the same animal are recovered in close proximity. This particular mammoth bone was recovered in 2010 with four other vertebrae. Photo credit: Tyler Kuhn.
Plate 9. A mammoth tusk exposed by placer mining near Dawson City, Yukon Territory, Canada. Although it took several days for the soil surrounding the tusk to thaw completely, we eventually recovered the entire 2.5 m, 45 kg tusk. The tusk is now part of the paleontological collection of the Department of Tourism and Culture in Whitehorse, Yukon Territory, Canada. Photo credit: Tyler Kuhn.
Plate 10. As running water cuts through the permafrost, the remains of ice organisms are exposed. Geologists believe that a small stream began to cut through this area of permafrost near the Yana River in northeastern Siberia around 60 years ago. When the cut reached an ancient lake, rapid erosion formed what is now the Batagaika crater. Such fresh exposures are common along rivers throughout Beringia. Photo credit: Love Dalén.
Plate 11. A first look at base camp. Ian Barnes and I pose for a photograph as the helicopter is unloaded on the Taimyr Peninsula in the Russian Far North. Other members of the 2008 expedition team have already donned their mosquito-net hats. Photo credit: Beth Shapiro.
Plate 12. Setting up and settling in. As the mosquitoes swarm overhead, the 2008 Taimyr expedition team begins to stake out its tent sites. Our site is at the top of a hill surrounded by lakes, all of which we will search over the next weeks for the remains of mammoths and other ice age animals. Photo credit: Beth Shapiro.
Plate 13. Another shot from the first day of our 2008 Taimyr expedition: my tent, and several million mosquitoes. Photo credit: Beth Shapiro.
Plate 14. An ice cave beneath the city of Yakutsk, Sakha Republic, Russia. Caves such as this one are often used in Siberian cities to store food during the summer months. At the far end of this ice cave, scientists prepare to display the Yukagir mammoth to a delegation of international scientists attending a conference. Photo credit: Beth Shapiro.
Plate 15. Wild Spanish ibex escape manipulation by the scientists leading the bucardo-cloning project. Accustomed to climbing vertical rock faces and balancing on narrow ledges, the wild ibex easily balance atop a thin ledge within the captive breeding facility, well out of reach of the research team. Photo credit: Alberto Fernández-Arias.
Plate 16. Grazed and ungrazed land in Sergey Zimov’s “Pleistocene Park” in the spring, after snowmelt. Ten years earlier, the area was a continuous community of willow shrubs. Today, the grazed area (foreground) in early spring has small amounts of green grass and freshly churned soil. This is caused by herbivores returning to this site during winter to graze and, in the process, trampling the snow and exposing the soil to the cold winter air. Photo credit: Sergey Zimov.