Neanderthal Man (9 page)

To give my frequent trips to Munich a veneer of science, I paid several visits to the Genetics Department of the Ludwig-Maximilians-Universität, where Barbara was a graduate student. Once, I even gave a seminar there on my experiments with ancient DNA. After my seminar the molecular biologist Herbert Jäckle asked me if I would be interested in an assistant professorship that would become available there in a couple of months. I said yes, seeing an opportunity to spend time on a more permanent basis with Barbara. But on a subsequent visit to Munich I realized that she had become involved with another scientist—one who studied fruit flies, as she did. Indeed, he would later become her husband. I flew back to Berkeley and did my best to forget about Barbara and Munich.

Six months later, I started to apply for jobs in earnest. I visited Cambridge University, where I was offered a lectureship; I visited Uppsala, where I was offered a position as a research assistant. Then, late one night, Germany caught up with me again, in the form of Charles David, the American-born dean of the biology faculty in Munich, who called me in Berkeley. Would I consider coming to Munich if the university offered me a full professorship instead of an assistant one?

This would amount to a tremendous advance in my career. Normally one would expect to be an assistant professor for a number of years before becoming a full professor. The full professorship was not just a title but came with resources such as a big laboratory, personnel, and funds. Yet I hesitated. I knew little of Germany, except for its reputation as home to one of the two worst political ideologies of the century. I had no idea whether I would fit in, or if my bisexuality would create problems. Ultimately, Charlie and Herbert together convinced me that Munich was a place where one could both live well and do science, so I decided to give it a try. My plan was to take the opportunity offered by the Munich professorship and do good work there for a few years so that I could then move back to Sweden. I accepted their offer and arrived in Munich with two big suitcases early one morning in January 1990, prepared to begin my independent scientific life in a world new to me and more than a little frightening.

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Setting up a laboratory is an intimidating experience, especially the first time you do it, even more so when you’re doing it in an unfamiliar environment. In my case, the environment was new in more than one sense. First of all, it was loaded with German history. The building where I was to work, the university’s Institute of Zoology, had been built and donated to the university by the Rockefeller Foundation during the Great Depression of the 1930s. During the war, it was bombed by the Americans, and it was rebuilt by the foundation after the war, so it epitomized the complicated and multifaceted relationship between Germany and the United States, a pendulum that has swung back and forth between the extremes of war and alliance. The institute was situated between the railway station and the complex of buildings erected by Hitler to house the Nazi Party headquarters. It was rumored that below the basement was a tunnel once used by the führer and his associates to move from the station to the headquarters. True or not, the rumor symbolized my fears of latent, subsurface German fascism.

Another novel aspect of the environment was that my appointment was to the Zoology Institute. I had never studied zoology, or even biology at the university level—just medicine, since in Sweden you can enter medical school directly after high school. This lapse became all too apparent almost as soon as I arrived, when an older professor asked whether I could perhaps teach the course on insect taxonomy in the upcoming semester. I was still jet-lagged, and preoccupied with other concerns, so without much thought I expressed my surprise that a zoology institute would deal with insects, as insects were hardly animals. In my mind, “animals” were things with paws, fur, and preferably floppy ears. The professor stared at me in disbelief and left without a word. I was immediately ashamed of having made such a complete fool of myself in the first week of my new job. But the good news was that no one ever again suggested that I teach any form of taxonomy or entomology at the institute.

As I was settling in, I learned that my predecessor at the institute had died unexpectedly of food poisoning. It was obviously not going to be easy to win the loyalty of all his colleagues, some of whom viewed me as an inexperienced and eccentric foreigner—a usurper of sorts. This was made clear in an unnerving encounter with Hansjochem Autrum, an emeritus professor and my predecessor’s mentor. Professor Autrum had been an influential figure in German zoology; when I arrived in Munich he still edited
Naturwissenschaften,
a somewhat influential German biology journal, and had an office on the same floor as my lab. During my first days in Munich, when I passed him on the staircase I greeted him cordially, but I got no response. One of my technicians reported that afterward he was heard to loudly complain that many good young German scientists could not find jobs and what did the department do but hire “international trash” (
internationaler Schrott
). I decided to ignore him from then on. Many years later, after his death, I became a member of a prestigious German order to which he had belonged, and I read his obituary in its proceedings. The author pointed out that before 1945 Professor Autrum had been a member not only of the Nazi Party but also of the Stormtroopers (SA) and had taught National Socialist ideology courses at a university in Berlin. Although I generally have a somewhat exaggerated desire to be liked by everybody, I felt retrospectively justified in having failed to befriend him.

Fortunately, Professor Autrum was an exception at the institute. Equally fortunately, he represented a generation that was on its way out in Germany. Gradually, by being frank about my ignorance not only of taxonomy but of most things zoological and administrative, I succeeded in bringing around even the older technicians in my group, and soon they wanted to help me build something that would be new and exciting. Charlie and Herbert, for their part, were extremely supportive. When the required laboratory renovations became more expensive than expected, the university came up with additional money. Slowly but surely, the equipment I needed was assembled, and all was put in order. Even more important, some students expressed interest in working with me.

Scientifically, I felt we needed to get systematic about establishing reliable procedures for amplifying ancient DNA. In Berkeley, I had begun to realize that contamination of these kinds of experiments with modern DNA was a serious problem, especially when the PCR was used. With the new PCR machines and the heat-resistant DNA polymerase, the process was  sensitive enough that under favorable circumstances a handful of DNA molecules, or perhaps even a single molecule, could start the reaction. That sounds wonderful, but could lead to trouble. If, for example, a museum specimen contained no surviving ancient DNA but a few DNA fragments from some museum curator, we could unwittingly wind up studying the curator’s DNA instead of the DNA of an ancient Egyptian priest. Extinct animals, of course, presented much less of an opportunity to mislead ourselves; in fact, it was in the course of doing such work that I first realized the huge potential for contamination, since sometimes when I tried to amplify mtDNA from animal remains, I would get human mtDNA sequences instead. In 1989, shortly before I left Berkeley for Munich, I had published a paper with Allan Wilson and Russell Higuchi, whose quagga work I replicated, in which we introduced what we called criteria of authenticity; these were procedures we thought had to be carried out before a DNA sequence retrieved by the PCR could be confirmed as truly old.
{13}
We recommended that a “blank extract”—that is, an extract with no ancient tissue but containing all other reagents to be used—be processed in parallel every time extractions from old specimens were performed. This allowed us to detect DNA that might lurk in the reagents themselves, which came to the lab from various suppliers. In addition, extractions and PCRs needed to be repeated several times, to ensure that a DNA sequence could be replicated at least twice. And finally, I had realized that hardly any fragments of ancient DNA were longer than 150 nucleotides. In short, I had concluded that many experiments purporting to have isolated ancient DNA that had been done up to that point, and especially before the PCR became available, were hopelessly naïve.

In hindsight, I now realized that the mummy sequence I had published in 1985 was suspiciously long given that my subsequent work had shown that ancient DNA was almost always degraded to small fragments. One of two factors could explain why the sequence I found, as another group demonstrated, came from a transplantation antigen gene
{14}
(precisely the type of genes we had studied in our lab back in Uppsala): either because I had identified the sequence with a probe for such genes or because a piece of DNA from the lab had contaminated my experiments. Given the length of the sequence, contamination seemed much more likely. I consoled myself with the thought that this is how science progresses: older experiments are overtaken by new and better ones. And I was happy to be the one to improve on my own work. With time, there also came help from outside the field. In 1993, Tomas Lindahl published a short comment in
Nature
in which he  suggested that criteria much like the ones we championed in 1989
{15}
were necessary for the ancient DNA field.
{16}
It was a great help to have a respected scientist from outside the field point this out—especially given my concern that the ancient DNA field tends to attract people without a firm background in molecular biology or biochemistry who, lured by the media attention that accompanies many ancient DNA results, simply apply the PCR to whatever old specimen they happen to be interested in. They practice what we in the lab liked to privately call “molecular biology without a license.”

As I now considered what projects to embark upon in my new lab, I was particularly inclined to study human history by molecular means. It was a fascinating topic but, as generally practiced, riddled with conjecture and biases stemming from preconceived ideas about history. I longed to bring a new rigor to the study of human history by investigating DNA sequence variation in ancient humans. One obvious possibility was to study the Bronze Age humans that were preserved in the peat bogs of Denmark and Northern Germany. But as I read more about them, I realized that these corpses had been preserved because the acid conditions in the bogs had essentially tanned them. Acid conditions lead to nucleotide loss and strand breakage and are therefore extremely bad for DNA preservation. But even worse, the tendency to find human DNA even in animal remains suggested that working with ancient humans could be seriously problematic.

So instead we started to collect samples of extinct animals, such as Siberian mammoths. And we started to do controlled experiments in a systematic way. For example, my first graduate students, Oliva Handt and Matthias Höss, used primers specific for human mtDNA. To my dismay, they found that they could amplify human DNA from almost all our animal samples and generally also from the blank extracts. We made up new reagents from fresh containers that had just been delivered to the lab, but it didn’t help. We did this again and again, trying to be as meticulous as we possibly could, but month after month we continued to find human DNA in almost every experiment. I began to despair. How could we ever trust the data, unless they completely conformed to our expectations, such as finding marsupial-like sequences from a marsupial wolf? And if we could only trust the expected results, that would make the field of ancient DNA very boring indeed, as we could then never discover the unexpected—which is, of course, the essence of experimental work and the dream of every scientist.

I walked home night after night frustrated and impatient with our failed experiments. But gradually it dawned on me that I was still being naïve about the contamination issue. I had not drawn the logical  conclusions from my awareness of the PCR’s extreme sensitivity. At Berkeley, and during the first period in Munich, we would extract DNA from museum specimens on our lab benches—the same benches where we handled large amounts of DNA from humans and other organisms we were interested in. If even a microscopic droplet of a modern DNA solution made it into the ancient DNA extract, the modern DNA would overwhelm the few ancient molecules that might have come from the ancient tissue. This could well happen even if we made no obvious mistakes, such as forgetting to change the plastic tip of a pipette.

Figure 4.1. Oliva and Matthias in the first “clean room” in Munich. Photo: University of Munich.

It became clear to me that what we needed was to achieve complete physical separation of the extraction and handling of DNA from ancient tissues and all other experiments in the lab. In particular, we needed to isolate these experiments from the PCR, where trillions of molecules were produced. We needed a laboratory dedicated solely to ancient DNA extraction and amplification. So we located a small windowless room on our floor, which we emptied out completely and repainted, then spent time thinking about how DNA that might be lurking on the new benches and instruments we bought for this lab could best be destroyed. We came up with some harsh treatments. We cleaned the entire lab with bleach, which oxidizes DNA. We mounted ultraviolet lamps in the ceiling and left them on all night, since UV light wreaks havoc on DNA molecules. And we bought new reagents for our new lab, the first “clean room” in the world devoted to work on ancient DNA (see Figure 4.1). These measures dramatically improved things. Our blank extracts became clean, while, to my delight, some of our samples continued to yield DNA. But gradually, over months, the blanks turned up with DNA again. I was furious. What was going on? We threw out all our reagents and bought new ones.