The Seven Daughters of Eve (2 page)

Robert runs the carbon-dating laboratory for archaeological samples in Oxford. He had been thinking about ways of getting more information from the bones that passed through his lab, aside from just dating them by the radiocarbon method. Collagen is the main protein not only in living bones but also in dead ones, and it is the carbon in the surviving collagen that is used to date them. Robert wondered if there was any genetic information in these surviving fragments of ancient collagen, so he and I put together a research proposal to study them. Collagen, being a protein, is made of units called amino-acids, arranged in a particular sequence. As we shall see in the next chapter, the sequence of amino-acids in collagen, and all other proteins for that matter, is dictated by the DNA sequence of their genes. We hoped to discover the DNA sequence of the ancient collagen genes indirectly by determining the order of amino-acids in the fragments of protein that survived in Robert's old bones. We advertised for research assistants several times but got no response at all. We would have expected a flood of applications for a regular genetics post, and put this zero interest down to the unusual nature of the project. Disappointingly few scientists want to venture from the mainstream field of research at an early stage of their careers. For us, this lack of a recruit meant we had to put back the start of the project by a year. Although very frustrating at the time, the delay proved to be a blessing in disguise – because, before the project got going, news came in of a new invention. A US scientist in California called Kary Mullis had dreamed up a way of amplifying tiny amounts of DNA – under perfect conditions, as little as a single molecule – in a test tube.

One warm Friday night in 1983 Mullis was driving along Highway 101 by the ocean; according to his account of events, ‘the night was saturated with moisture and the scent of flowering buckeye'. As he drove, he was talking to his girlfriend, seated beside him, about some of the ideas he had been pondering to do with his work at a local biotech company. Like everyone else in the genetic engineering business, he was making copies of DNA in test tubes. This was a slow process because the molecules had to be copied one at a time. DNA is like a long piece of string, and the copying started at one end and finished at the other. Then it started at the beginning again and you got another copy. He was talking out loud about this and suddenly realized that if, instead of starting the copying at one end only, you started at
both
ends you would start what would effectively be a sustainable chain reaction. You would no longer just be making copies of the original but copies of copies, doubling the number at every cycle. Now, instead of two copies after two cycles and three copies after three cycles, you would double up after each cycle, producing two, four, eight, sixteen, thirty-two, sixty-four copies in six cycles instead of one, two, three, four, five and six. After twenty cycles you would have not just twenty copies but a million. It was a real ‘Eureka' moment. He turned to his girlfriend to get her reaction. She had fallen asleep.

This invention, for which Kary Mullis rightly won the Nobel Prize for Chemistry in 1993, genuinely revolutionized the practice of genetics. It meant that you could now get an unlimited amount of DNA to work on from even the tiniest piece of tissue. A single hair or even a single cell was now all that was needed to produce as much DNA as you could ever want. The impact of Mullis's brainwave on our bone project was simply that I decided to forget about working on the collagen protein, which would have been horrendously difficult, and use the newly invented chain reaction to amplify what, if anything, was left of the DNA in the ancient bones. If it worked, then we would get vastly more information from the DNA than we would ever have got from the collagen. We would be going directly for the DNA sequence itself, rather than inferring it from the amino-acids. Much more importantly, we would be able to study
any
gene, not just the ones that controlled collagen.

At last we got an answer to our advertisement for a research assistant, and Erika Hagelberg joined the team. We were obviously not going to get anyone with previous experience in working with ancient DNA, because it had never been done before, but Erika's degree in biochemistry, combined with research posts in homoeopathy and in the history of medicine, reflected a combination of a solid scientific training and the catholic interests which suited the project. Besides, she was the only applicant. Now we needed some very old bones.

News came in during 1988 of an excavation going on in Abingdon, a few miles south of Oxford. A new supermarket was going up and the mechanical diggers had ploughed into a medieval cemetery. The local archaeology service had been given two months to excavate the site before the developers moved back in, so when Erika and I arrived, it was buzzing with activity. It was a hot and brilliantly sunny day and dozens of field assistants, stripped down to the bare essentials, were dotted all round the site scraping at the earth with trowels, rummaging around in deep pits or wading through water-filled trenches. Several skeletons lay half-exposed, encrusted with orange-brown earth, criss-crossed by strings which marked out a reference grid. As we gazed down at them, our prospects didn't look at all promising. Having worked with DNA for several years, I was trained to treat it with respect. DNA samples were always stored frozen at 70° below zero, and whenever you took DNA out of the freezer you were taught always to keep it in an ice bucket. If you forgot about it and the ice thawed then you had to throw the DNA out because, so everyone assumed, it would have degraded and been destroyed. No-one imagined it would last for more than a few minutes on the laboratory bench at room temperature, let alone buried underground for hundreds or even thousands of years.

Nevertheless, it was worth a try. We were allowed to take three thigh bones from the excavation away with us. Back in the lab we had to make two decisions: how to get the DNA out, and what section of DNA to choose for the amplification reaction. The first was easy enough. We knew that if there were any DNA left at all it would probably be bound up with a bone mineral called hydroxyapatite. This form of calcium had been used in the past to absorb DNA during the purification process, so it seemed quite likely that the DNA would be stuck to the hydroxyapatite in the old bones. If that was the case, we had to think of a way of disengaging the DNA from the calcium.

We cut out small segments of bone with a hacksaw, froze them in liquid nitrogen, smashed them up into a powder, then soaked the powder in a chemical which slowly took out the calcium over several days. Fortunately, when all the calcium had been removed, there was still something left at the bottom of the tube – a sort of grey sludge. We guessed this was the remnants of the collagen and other proteins, bits of cells, maybe some fat – and, we hoped, a few molecules of DNA. We decided to get rid of the protein using an enzyme. Enzymes are the catalysts of biology, making things happen much more quickly than they otherwise would. We chose an enzyme which digests protein, rather like the ones in a biological washing powder which get rid of blood and other stains for the same reason. Then we got rid of the fat with chloroform. We cleaned what was left with phenol, a revolting liquid which is the base for carbolic soap. Even though phenol and chloroform are both brutal chemicals, we knew they did not harm DNA. What remained was a teaspoonful of pale brown fluid which, theoretically at least, should contain the DNA – if there was any. There would be at best only a few molecules, so we had to use the new DNA amplification reaction to boost the yield before we could carry out the next steps.

The essence of the amplification reaction is to adapt the system for copying DNA that cells use. Into the tube go the raw materials for DNA construction. First in is another enzyme, this time one used for copying DNA; it is called a polymerase and gives the reaction its scientific name – the
polymerase chain reaction
or PCR for short. Next, a couple of short DNA fragments are added to direct the polymerase enzyme to the segment of the original DNA that is to be amplified and ignore everything else. Finally, the raw materials – the nucleotide bases – for building new DNA molecules go into the mix along with a few ingredients, like magnesium, to help things along. Plus, of course, the stuff you want to amplify – in our case, an extract of the Abingdon bone containing, we hoped, a few molecules of very old DNA.

Then we had to decide which gene to amplify. Because we knew there wasn't going to be much, if any, DNA left in the bone extract we decided to maximize our chances by choosing something called mitochondrial DNA. We chose mitochondrial DNA for the simple reason that cells have upwards of a hundred times more of it than any other gene. As we will see, mitochondrial DNA turns out to have special properties which make it absolutely ideal for reconstructing the past; but in the first instance, we chose it as our target simply because there was so much more of it than any other type of DNA. If there was any DNA at all left in the Abingdon bones, then our best chance of finding it was by targeting mitochondrial DNA.

So, into the reaction went all the ingredients necessary for amplifying mitochondrial DNA, plus a few drops of the precious bone extract. To get the reaction to fire in the tube you need to boil it, cool it, warm it up for a couple of minutes; then boil it again, cool it, warm it up…and go on repeating this cycle at least twenty times. Modern genetics laboratories are full of machines for doing this reaction automatically. But not then. Back in the 1980s the only machine on the market cost a fortune, and there was no money for one in our budget. The only way to do the reaction was to sit with a stop-watch in front of three water baths, one boiling, one cold and one warm, and move the test tube by hand from one bath to the next every three minutes. Then do it again. And again. For three and a half hours. I only tried it once. The reaction didn't work and I was bored stiff. There had to be a better way. What about using an electric kettle? I spent the next three weeks with wires, timers, thermostats, relays, copper tubing, a washing-machine valve and my kettle from home. In the end I had a device that did all the right things. It boiled. It cooled (very fast) when the washing-machine valve opened and let cold tap-water into the coils of copper tubing. And it warmed up. And it worked.

We could see that the machine (christened the ‘Genesmaid', after the tea-making device people of a certain age regard as an essential bedroom accessory) had managed to get the amplification reaction to work not only with a control experiment using modern DNA but also, very faintly, with the Abingdon bone extract. By comparing its sequence to those published in scientific papers, it didn't take us long to prove that the DNA was genuinely human. We had done it. Here, in front of our very eyes, was the DNA of someone who had died hundreds of years ago. It was DNA resurrected, literally, from the grave.

Now, looking back, it is hard for me to believe that the research set in motion by the recovery of DNA from those crumbling bones in the Abingdon cemetery, the bones which looked so unpromising when I first saw them half-buried in the earth, should lead over the following years to such profound conclusions about the history and soul of our species. As my story unfolds you will see that, like most scientific research, this was not a seamless progression towards a well-defined goal. It was more like a series of short hops, each driven as much by opportunity, personal relationships, financial necessity and even physical injury as by any rational strategy. There was no set path towards the discovery of the Seven Daughters of Eve. The research just moved a little bit at a time, mostly forwards, towards the next dimly visible goal, informed by what had gone before but ignorant of what lay ahead.

At the time, though our result was a great triumph, strangely enough it didn't feel like it. I think Erika and I were too heavily involved in the details to appreciate the significance of what we had achieved. Besides, by then we were not getting on at all well. Tension had been building for weeks because, for some reason, Erika and I did not seem to be working together effectively. Only much later did I start to realize what our breakthrough could mean, not only for science but for popular history as well. That would come later; at the moment we had more pressing claims on our attention. I had heard on the grapevine that other research teams were also looking for DNA in old bones. This meant we had to get our work published with maximum speed, otherwise there was a real danger that we would be scooped. What counts in science is not being the first to do an experiment but being the first to publish the results. If someone else published even a day before we did, then they would claim the prize. Fortunately, the editor of the scientific journal
Nature
was persuaded to rush our paper into print in record time, and it was published just before Christmas 1989.

I was quite unprepared for what happened next. Although my previous research on brittle bone disease had occasionally been covered in the local papers and even once or twice in the nationals, it could not be said that any new result had sparked off a media frenzy. So it was a new experience when I got into work next day to find the phone constantly ringing with press enquiries. A few years previously I had actually spent three months in London as a reporter for ITN, which runs the television news service for the main commercial terrestrial channels in the UK. This venture was part of a well-intentioned fellowship scheme run by the Royal Society, designed to bridge the gap between science and the media. I was attracted to it by the generous expenses with which I hoped to pay off my bank overdraft. In fact, I ended up owing more money than I had to start with, not least because of the amount of time I spent in bars and restaurants with the well-heeled professionals. One night, for instance, I was precocious enough to offer to buy a drink for one well-known presenter. ‘Thanks, dear boy, I'll have a bottle of Bollinger,' came the great man's answer. What could I do but comply? Still, though a financial disaster of major proportions, those few months taught me many things about the news media, including the way to trim my replies to reporters' questions down to the simple sentences I knew they wanted.