Warning signs

Potential problems with the samples were apparent in what were likely the first experiments done with the DNA isolated from them. These were amplifications of specific human DNA sequences using a technique called the polymerase chain reaction, or PCR. By using short DNA sequences that match parts of the human genome, it's possible to start with a single DNA molecule and create many copies of it, which makes it simple to detect its presence. In this case, the PCR reactions targeted sequences that are known to vary in length in the human population—a feature that makes them useful for forensic identification.

If the DNA was human and had not degraded much during its time in the environment, then most of these reactions should produce a clear, human-like signal. The same would be true if, as Ketchum concluded, the samples contained DNA from a close relative of humans (remember, chimps' DNA is over 95 percent identical to ours). If the animal were more distantly related, you might expect some reactions to work and some to fail, with the percentage of failures going up as the degree of relatedness fell. In some cases, you might expect the reactions to produce a PCR product that was the wrong size due to changes in DNA content that occur during evolution.

But you can't necessarily expect the DNA to sit outdoors and remain intact. DNA tends to break into fragments, with the size of the fragments shrinking over time. Depending on how degraded the sample is, you might see more or fewer reactions failing.

What they saw was a chaotic mix of things. As Ketchum herself put it, "We would get these crazy different variants of sequence." Some reactions produced the expected human-sized PCR products. Others produced products with unexpected sizes. Still others produced the sorts of things you'd expect to see if the PCR had failed entirely or there was no DNA present. "We would get these things that were novel in genbank. We would get a lot of failure, and we'd get some that would have regular human sequence," Ketchum said. "We could not account for this, and it was repeatable."

All of which suggested that there was likely to be DNA present that was only distantly related to humans; anything that was from a human or close relative was probably seriously degraded.

In fact, the team did an experiment that suggested this was exactly what they were dealing with: they imaged the DNA using electron microscopy. This revealed exactly what their initial experiments suggested: shorter fragments of DNA, some of it a single (rather than double) helix. Strands that paired nicely for some stretches and then diverged into single stranded sections, which then paired again to a completely separate molecule. This sort of pattern is what you might see if there were some distantly related mammals present, where the protein-coding sequences would match fairly well, but the intervening sequences would probably be very different.

So all the initial data suggested that the DNA was badly preserved and probably contaminated. Which in turn suggests that whatever techniques they used to get DNA from a single, uncontaminated source just wasn't sufficient for the samples they were working with. But instead of reaching that conclusion, the bigfoot team had an alternative: their technique worked perfectly fine. It was the sample that was unusual.

The problem is that it simply couldn't be that unusual. The idea is that there was some other primate that was still capable of interbreeding with humans. In the cases where we know this happened (semi-modern humans like Neanderthals and Denisovans), the DNA sequences are so similar that it's quite hard to tell them apart. Here, the team was seeing indications that human DNA was mixed with something that was really quite distant—probably not even one of the great apes.

These were far from the last results that should have told them they were on the wrong track.

Looking suspiciously human

Nevertheless, the authors plowed on. And one of the first things they found was that at least some of the DNA was human. This, as it turned out, was the foundation for their conclusion that the DNA was from a human-primate hybrid.

It's often overlooked that human cells actually have two genomes. One lives in the chromosomes stored in the nucleus, and that's the one we're typically concerned with. But a second resides in our mitochondria, small compartments in the cell that provide most of the cell's ATP. These are the remains of what were once free-living bacteria but took up a symbiotic residence inside the cell billions of years ago; however, they still have a small genome of their own (circular, like bacteria's) with a handful of essential genes on it.

There are a few things that make mitochondrial DNA effective for tracking populations of humans and other species. Because this genome doesn't have a full DNA repair machinery at hand, and because it can't undergo recombination, it tends to pick up mutations far more rapidly than the nuclear genome. That means that even closely related populations are likely to have some differences in their mitochondrial DNA. There are also hundreds of mitochondria in each cell, and each of these may have dozens of copies of the genome. So it's relatively easy to get samples, even from badly degraded and/or contaminated DNA like that found in ancient bones.

So team bigfoot sequenced the mitochondrial genome of several of their samples. And rather than a novel primate sequence that was distantly related to humans, the sequences were human. Which is what you might expect if the species is a hybrid as the authors concluded. What you wouldn't expect is that the sequences would come from multiple humans—from the wrong side of the planet.

All indications are that successful interbreeding between humans and closely related groups like Neanderthals and Denisovans was relatively rare. You'd expect that something that looks like a walking shag carpet would be more distantly related, and that it would be much, much harder to successfully interbreed. This makes the hybrids even rarer. Instead, each sample tested produced a different mitochondrial DNA sequence, which implies the interbreeding had to have taken place many, many times. (And that the hybrids never bred with females of whatever the primate in question was. And that said primate is, apparently, extinct, since none of its mitochondrial DNA showed up.)

Who were these human females that ostensibly did the interbreeding? If you wanted to make a scientifically plausible guess, you'd bet on the mitochondrial DNA lineages that originate in Asia (most likely those branches that expanded into the Americas). Those are the only humans that are likely to have been around until a few hundred years ago. And that's exactly what they didn't find. Instead, most of the sequences originated in the human populations of Europe, with an African sample or two.

And at least one of them was recent—Ketchum described one of the mitochondrial sequences in detail, saying, "about 13000 years ago is when that haplotype came into existence. It was in Spain, basically, where it originated. So the hybridization could not have occurred before that haplotype came into existence." In her view, that put an upper limit on when these sequences made it to North America. "It couldn't have been longer than 13,000 years ago," she told Ars.

On the face of it, there's simply no way to make sense of this—the European and African DNA, the recent time frame for its arrival, the fact that there must have been so many interbreedings.... The obvious interpretation is that the samples were all from humans or contaminated with human DNA, which nicely explains the diversity and modernity of the sequences.

But remember, to Ketchum, that possibility had been ruled out. In the absence of the obvious, her team went with a far less obvious suggestion: sometime during the last glacial period, a diverse group of Europeans and Africans got together and wandered across the vast empty spaces of the Greenland ice sheet and found themselves in North America. "Several of the Smithsonian scientists even wrote a book about it, where they've gone below the Clovis layer and found artifacts that they feel came from [an] area in France," she said. But she wasn't committed to that idea and later suggested that the interbreeding might have taken place in Europe... after which the Sasquatch left to cross the Bering Sea-land bridge before the Ice Age ended. "It's feasible they could have crossed the world, basically," she said. "They're very fast."

Ultimately, though, Ketchum indicated these are just technical details. She wasn't especially interested in sorting them out. "We don't know how they got here, we just know they did."