The seventeenth paper presented at the July 1992 conference in Ekaterinburg was given by a cheerful, dark-haired, forty-one-year-old molecular biologist, Dr. Pavel Ivanov of the Englehardt Institute of Molecular Biology of the Russian Academy of Sciences in Moscow. Ivanov’s subject was DNA testing. He told the conference that, at the end of 1991, Russian Chief Medical Examiner Vladislav Plaksin had asked him to look into the possibility of using this new technique to help identify the bones found by Alexander Avdonin and Geli Ryabov. Ivanov knew that this work could not be done in Russia. No one “in Russia had the experience of working with bone materials,” he explained to the conference, nor did Russia possess the necessary technology. Nevertheless, while in London in December 1991, he had visited what he called the “Central Criminal Research Center” of the British Home Office at Aldermaston in Berkshire and begun negotiations for a joint British-Russian study of the bones.
Early in July 1992, just two weeks before this conference, an agreement had been reached between the Russian Ministry of Health and the British Home Office. A joint effort would take place at Aldermaston involving Dr. Peter Gill, director of the Molecular Research Center of the Home Office Forensic Science Service, Sir Alec Jeffreys of Leicester University, founder of the DNA fingerprinting technique, and Dr. Erika Hagelberg of Cambridge University, a specialist on molecular genetic analysis of bone remains. The Russian scientist would be Ivanov himself. All expenses except travel would be borne by the British Forensic Science Service, and travel costs (essentially Ivanov’s airfare to and from Britain) would be paid by the Sverdlovsk regional government, which had approved the arrangement. The tests in England, Ivanov told his audience, would enable the researchers to determine whether, among the nine exhumed skeletons, a family group existed. Further, if enough uncontaminated DNA could be extracted from the remains, and if living persons descended from blood relations of the Imperial family could be persuaded to donate samples for comparative purposes, it would be possible to prove whether or not the family group found in the grave was that of Tsar Nicholas II.
The fact that the bones were going to England because DNA technology was not available in their own country was embarrassing for Russian scientists. “We were working on molecular genetic testing at one time,” said Nikolai Nevolin with a wry smile. “Academician Vavilov began using this method. Then Mr. Stalin shot his entire team. As a result, we began lagging behind.”
When Stalin died in 1953, Pavel Ivanov was two years old. Twenty years later, in the era of Brezhnev, Ivanov was on his way to acquiring a degree in molecular biology at Moscow State University—“the best we have in Russia, very well regarded in Europe.” He began as a pure research scientist, working on the international human genome project at the Institute of Molecular Biology. In 1987, his group, trying to read the genetic codes which create human beings, discovered a technique of DNA fingerprinting similar, but not identical, to work first done by Alec Jeffreys in England. Ivanov, still a basic scientist, began to explore and develop this technique. His work came to the attention of “everyday, working” organizations, such as the forensic crime laboratory and the KGB. “They expressed interest in practical applications of my work and suggested that I establish a forensic DNA laboratory,” he explained. “I agreed because forensic science was very interesting for me and because, with the level of crime so high in Moscow, I thought I might be able to do something to help. I did not work for the KGB; I have never been a Communist. But I understood the potential of these techniques for combating crime. Since then, I have had two jobs. I have kept my position as a pure scientist at the Institute of Molecular Biology and I have also become the DNA adviser to the chief medical examiner of Russia, Dr. Plaksin. Later, when the Romanov case arose, I became the principal Russian DNA investigator by appointment of the Russian state procurator general.”
Ivanov worked in both places in order to earn money. His wife, an assistant professor of biology, had a small salary; he helped his mother, a retired economist with a miserably inadequate pension; and he had two children. Despite his workload, he considered himself fortunate. He traveled far more widely than most Russian scientists, attending conferences as far afield as Australia and Dubai. He had worked at the FBI DNA laboratory in Washington, D.C., and had traveled back and forth across the United States. In the mid-1990s, his work on the Romanov bones had made him Russia’s best-known molecular biologist. During the summer of 1994, he drove his Volvo from Moscow to Ulm on the Danube in southern Germany in order to perform DNA tests on the remains of a recently deceased Russian emigre who claimed to have been the Tsarevich Alexis. During a long evening in a German restaurant, he talked about his involvement with the Romanov bones:
“I was the one who decided we should go to England when Plaksin asked for my recommendation,” Ivanov said. “The FBI laboratory in Washington and the AFIP both do excellent DNA work, but I chose Peter Gill because I knew him and because the British Forensic Science Service had the highest level of expertise in this particular sort of investigation—that is, using mitochondrial DNA. Also, of course, I already had considered asking Prince Philip, the Duke of Edinburgh, to help us. And I knew that he would be much more likely to help if the work was being done in England. But we had to find money. For a Russian scientist today, it is always a question of money. There are no political barriers, but there are financial barriers. You can’t go where you want.”
On September 15, 1992, Pavel Ivanov boarded a jet in Moscow. With him in a British Airways travel bag, carefully wrapped and sealed in polyethylene, he carried pieces of the femurs of each of the nine skeletons lying on tables in the Ekaterinburg morgue. At Heathrow, Ivanov was met by Nigel McCrery, a BBC television producer who had been active in the negotiations to bring the bones to England.* McCrery, feeling that it was “inappropriate to carry the Russian Imperial family in the boot of my Volvo,” had hired a Bentley limousine from Co-operative Funeral Services. In style, therefore, Ivanov, McCrery, and the Romanov remains were driven to Peter Gill’s house in the woods near Aldermaston, where the three men took photographs to memorialize the occasion. In the morning, Gill and Ivanov carried the bones through the high barbed-wire fences and security checkpoints of Britain’s huge Ministry of Defence atomic-research facility at Aldermaston. Inside the complex and off to one side, the Forensic Science Service had been given a small building to use as a research laboratory. For the next ten months, the two men and a team of others would attempt to compare and match the DNA of the Ekaterinburg skeletons with one another and with that of living relatives of the murdered Russian Imperial family.
If the Central Crime Laboratory of the U.S. Federal Bureau of Investigation were to be turned into a private business, instructed to develop a “commercial, customer-focused, cost-conscious approach,” and told to break even or make a profit by charging fees and offering its services to all who walk in the door, this would approximate what happened recently to Britain’s equivalent of the FBI lab, the Home Office Forensic Science Service. For fifty years, beginning in the 1930s, the Forensic Science Service functioned as a source of expert investigative assistance to provincial and local police departments in England and Wales. Experts from FSS scrutinized evidence in cases of murder, rape, arson, burglary, drug use, poisoning, and forgery. They visited crime scenes, examined bodies, fingerprints, weapons, bullets, stains, alcohol levels, handwriting samples, and old typewriters. The beneficiaries of this specialized knowledge were the Crown prosecutors, on whose behalf the FSS provided expert testimony in court. The people of Britain paid for these services by paying taxes.
In April 1991, the FSS was overtaken by Thatcherism. Its six hundred scientists, technicians, and other staffers, working at six laboratories scattered around the country, suddenly were transformed into scientific guns for hire. The FSS became a business, required to make its own way in the world by charging fees for services. The door was opened to everyone—“widening the customer base” was the terminology used. Defense attorneys, foreign governments, insurance adjusters, regional health authorities, and private citizens were invited in. The transformation was “turbulent,” admitted FSS Director General Janet Thompson. To most scientists, “the world of business still seemed radical.” In 1991–92, police case work dropped 18 percent and a deficit of £1.1 million was posted. But the following year, matters improved. The police came back and paid the required fees. The FSS made a profit equal to its earlier loss. Most spectacularly, in the summer of that year, the service and its leading molecular biologist, Dr. Peter Gill, made headline news, not only in Britain but around the world.
Dr. Gill, the head of Biological Services (Research) of the Forensic Science Service, is a slightly built man in his early forties, about five feet nine, with a pale face, uncombed hair, a brown mustache, and watchful eyes behind thick spectacles. He owns a dark blue suit, which he wears for press conferences, but in and around his laboratory his typical dress is a tattered sweater, shapeless corduroys, and elderly loafers. Born in Essex, Gill did an undergraduate degree in zoology at Bristol University, received a doctorate in genetics from Liverpool University, and did a five-year postdoctoral fellowship in genetics at Nottingham University. In 1982, he joined the Forensic Science Service Research Laboratory at Aldermaston to work on forensic applications of conventional blood typing methods. In 1985, against strong opposition in the service, he began to study the utility of DNA profiling in forensic science. Aware of the significance of Alec Jeffreys’s work, he briefly joined Jeffreys’s laboratory and, that same year, coauthored with Jeffreys the first scientific paper which demonstrated that DNA profiling could be used in forensic science. The methods described in this paper are now routinely used around the world. Gill himself has published over seventy papers in the scientific literature.
Although he is shy and speaks cautiously with strangers, there is one point on which Dr. Gill is quietly emphatic: his laboratory is the best of its kind in the world—“We have retained our world lead” is his way of putting it. In his opinion, therefore, it was entirely understandable that Pavel Ivanov had wished to bring the Russian bones to Aldermaston. “Ivanov asked me a long time ago whether we’d be interested in carrying out these tests,” Gill said. “When he asked, I had to go through the Home Office. They considered all the political ramifications, and eventually we got the go-ahead.”
The political ramifications existed on many levels. The most obvious was the current relationship between John Major’s Conservative government in Britain and Boris Yeltsin’s presidency of Russia. Both parties were interested in bringing to fruition a long-suspended diplomatic project: a visit to Russia by the queen. No British monarch had visited Russia since 1908, when King Edward VII and Queen Alexandra came by yacht to Tallinn (then Reval) to visit Tsar Nicholas II and Empress Alexandra.* Mikhail Gorbachev and Boris Yeltsin both had invited the queen to come, and Her Majesty and the British Foreign Office wanted the visit to take place.
But first there was some unfinished historical and family business. The Russian Imperial family and the British Royal family were closely related. King George V, Elizabeth II’s grandfather, was Nicholas II’s first cousin. Indeed, so close was the physical resemblance between the cousins that at George’s wedding, Nicholas often was mistaken for the groom. King George also was a first cousin of Empress Alexandra. In the spring of 1917, after the tsar had abdicated and while Alexander Kerensky and the Provisional Russian government were trying to provide for the safety of the Imperial family by sending them to political asylum abroad, King George V at first welcomed a proposal that Britain bring his Russian cousins to safety by ship. Then the king—fearing that the former tsar’s unpopularity in Britain would tarnish the British monarchy—reversed himself and insisted that they not be brought. George V’s act helped doom Nicholas, his wife, and his five children. When the British door slammed shut, Kerensky sent the family to Siberia, hoping to put them out of reach of the Bolsheviks. They still were there when, seven months after Kerensky’s fall, Lenin’s long arm reached out.
This catastrophe led to many recriminations. Members of the Russian Imperial family who escaped, aristocratic emigres, and numerous White Russians abroad bitterly condemned King George and his family and descendants. For three quarters of a century, many Russians have regarded England with deep suspicion and resentment. The British Royal family is aware of this hostility. Over the years, palace officials have attempted to bury the king’s role in the Romanov tragedy; official biographers of George V were advised that they should “omit things and incidents which were discreditable.” In 1992, the possibility that the Romanov bones might come to Britain to be verified by British scientists with the help of British Royal persons offered an opportunity to put some of these passionate feelings to rest.
According to a Forensic Science Service spokesperson who stays close to Dr. Gill specifically to answer nonscientific questions, the decision to bring the bones to Aldermaston was made on a relatively low level; that is, by Janet Thompson, the FSS director general. “Of course,” said the spokesperson, “with the high profile that came with this project, we put it before the home secretary. He could have objected if he had wanted to.” The spokesperson does not know whether Kenneth Clark discussed the project with the foreign secretary or the prime minister. Or whether anyone thought to consult the Royal family. If this was not done, however, Dr. Thompson and Secretary Clark were assuming historical and diplomatic responsibilities far beyond the normal range of their professional and political assignments.
There was one area in which Thompson—no doubt supported by Clark—did make a decision on her own. This was the decision to ignore the new Thatcherite decree that the FSS was to charge for services and attempt to make a profit. The service spent a large sum of money on the Romanov project. “We did all nine of them, the whole lot,” said Peter Gill. “It managed to be expensive.” “It was very expensive,” chimed in the spokesperson, adding that no figure is available. The sum can be roughly estimated. A year later, the FSS negotiated with a private citizen to perform DNA testing on an unknown woman and a possible relative. These tests were to be performed on preserved tissue and recently drawn blood, both sources from which DNA is far easier to obtain than from old, long-buried bones. For this work, the FSS demanded a five-thousand-pound down payment, plus another five thousand pounds placed in escrow in an English bank. All of this money was spent. The Romanov project involved typing and comparing bone fragments from nine people in Russia, plus blood samples from at least three relatives alive today. Even using the same expense figures for much more difficult tests, this would mean that twelve DNA profiles would cost sixty thousand pounds (over $100,000). Dr. Alka Mansukhani, an American molecular biologist routinely doing DNA extraction and sequencing at New York University Medical Center, believes that, if overhead was included, the figure probably is accurate.
The Home Office and FSS accountants funded these costs as pure research.
An adult human body is a cohesive mass of 80 trillion cells, yet in all this amplitude and diversity, there is an extraordinary sameness: each one of these cells contains all the genetic information needed to produce a complete and unique human being. This hereditary knowledge is carried in the chromosomes; in a normal person, forty-six in each cell nucleus, twenty-three from the mother, twenty-three from the father. Chromosomes are made up of molecules of DNA (deoxyribonucleic acid), which use their own chemical structure to store genetic information and commands. The DNA molecules are created from four basic chemical building blocks called bases, and the sequence in which these bases occur provides the information necessary to commence and control the building of a human body. For simplicity’s sake, molecular biologists describe the four bases by their initial letters, A, G, C, and T (adenine, guanine, cytosine, and thymine). The bases appear in pairs bonded with hydrogen; A bonds with T; G bonds with C; these combinations are known as base pairs. In 1953, James Watson and Francis Crick discovered the detailed, overall molecular structure of DNA. They found long, tightly coiled strands, each resembling a ladder twisted into the form of a spiral staircase. The A, C, G, and T base pairs formed the rungs; the sides of the ladder, to which the rungs were attached, were made up of alternating molecules of sugar and phosphate. Watson and Crick named their discovery the double helix.
The unique structure of every individual human body is dictated by the different combinations of these four letters in base pairs in the DNA. For example, at some point in the strand, one individual will read A, C, G, T, C, C, T. Another person, in the same part of the strand, will display a different sequence, say A, T, T, C, A, G, C. Whatever the base pair sequence, each cell in a human body contains the same DNA sequence, storing the same information and commands. But, to avoid massive confusion, nature activates only that part of the command system necessary for the function of that particular cell.
Each cell with its set of forty-six chromosomes contains approximately 3.3 billion DNA base pairs, strung together in clusters of spiraling double helixes. If one magnified this structure to a humanly visible five characters (A, G, T, C, T) per half inch, it would require a strip of paper 162 miles long to write the entire base sequence of a single chromosome. Approximately 99.9 percent of the 3.3 billion base pairs found in a single cell appear in the same sequence in all human beings; they ensure that all humans possess similar characteristics: two eyes, two ears, one nose, ten toes, blood, saliva, stomach acid, and so on. However, in the remaining 0.1 percent (that is, 3.3 million base pairs), the sequence of these base pairs differs from one person to another. It is the fact that individuals vary at this basic molecular level that now permits scientists to determine which human being was the source of this or that sample of bone or tissue, blood, semen, or saliva.
In the early 1980s, Dr. Alec Jeffreys, working at Leicester University, first recognized the enormous potential of the DNA variable in human beings to resolve questions of identity. He identified regions within hypervariable areas and used radioactive isotopes, called probes, to create an image on film of the DNA strands extracted from individuals. These visible symbols appear strikingly similar to the bar codes which are printed on packages and cans at every supermarket. The DNA patterns—Jeffreys called them “DNA fingerprints”—could then be used to compare one person’s DNA with that of another. Because children would derive half of their DNA base pairs from their mother and half from their father, family relationships could be established or refuted. In 1983, a boy was refused entry into England because an immigration officer doubted that he was the son of a Ghanaian woman who had rights to settlement in the United Kingdom. Jeffreys’s new DNA technique was employed and proved that the boy was the woman’s son. The chance of this match occurring at random was one in ten million.
Within less than a decade, DNA typing has become the most powerful forensic science tool since the nineteenth-century discovery that the fingerprints of no two persons are the same. Comparisons of DNA now routinely solve paternity cases. Murderers are identified by samples of blood, hair, other tissues, or fluids, liquid or dried. Samples of DNA from bones and teeth have helped resolve long-standing mysteries involving missing persons and unidentified bodies. DNA is remarkably stable: it has been extracted and identified from a three-thousand-year-old Egyptian mummy, from a seven-thousand-year-old mammoth, from the dried saliva remaining on a licked postage stamp. And, properly handled and identified, it is unerring. No prosecutor or defense attorney, no historian, no churchman of any faith, no believer in any political ideology, can disprove the essential message of DNA: that every human being is distinct from every other. DNA evidence, declared one American district attorney, is “like the finger of God pointing at someone and saying, ‘You are the one!’ ”
Because of the age and deteriorated condition of the Romanov bones, Dr. Gill and Dr. Ivanov faced a task radically more difficult than in any previous DNA typing examination. In a sterile environment, they began by grinding away one millimeter of the contaminated outer surfaces of the bones with sand wheels attached to a high-speed electric drill. The remaining bone was frozen in liquid nitrogen, then pulverized to a fine powder and dissolved in various solutions, then centrifuged to release a microscopic quantity of DNA. So paltry and degraded, in fact, were the sample yields that Gill and Ivanov applied an even more recently developed technique called PCR (polymerase chain reaction), in which selected relevant sections of base pair strands are chemically duplicated over and over in a test tube to provide sufficient quantities of DNA material for scientists to study.
Using nuclear DNA, the Aldermaston team first turned to determining the sex of each of the skeletons. A gene on the X chromosome (females have two X’s) is six base pairs longer than the similar gene on the Y (males have one X and one Y) chromosome. Using PCR, the scientists could obtain sufficient material to measure and determine this six-base-pair difference. The result was a confirmation of the anthropological findings of Abramov and Maples: there were four males and five females. Next, still using nuclear DNA and studying base pair sequences, Gill and Ivanov tested all nine for a family relationship. Short tandem repeat (called STR.) sequences are natural base pair repetitions in certain hypervariable regions of a chromosome—say, T, A, T, T—occurring over and over again. Within a family, these sequences and the number of repetitions tend to be constant; a different sequence or different number of repetitions in each individual sample would indicate that no family group was present. Again, the results were what was to be expected if the bones had come from the Imperial party. In Gill’s words: “Skeletons 3 through 7 exhibited patterns which would be expected in a family group where 4 and 7 were the parents of children 3, 5, and 6.” The other four adults were excluded as possible parents. Further, Gill’s report continued, “If these remains are the Romanovs then … test data indicated that one of the daughters and the Tsarevich Alexis were missing from the grave.” Other tests established paternity. STR DNA patterns from Body No. 4 were found in No. 3, No. 5, and No. 6; thus, the adult male presumed to be Nicholas was confirmed to be the father of the three young women. This was as far as Gill and Ivanov could go using the small quantity of degraded nuclear DNA available. They had established a party of four males and five females. They had established a family: a father, a mother, their three daughters. But to identify these men and women—to give them names—they had to try another tack.
Fortunately, a second form of DNA is available in human cells. Called mitochondrial DNA, it appears plentifully in units outside the nucleus which function as power stations for the cell. Mitochondrial DNA is inherited independently of nuclear DNA, and whereas nuclear DNA is inherited half from the mother, half from the father, mitochondrial DNA is inherited exclusively from the mother. From mother to daughter, it is transmitted intact, “passing from generation to generation unchanging, like a time machine,” said Gill. “The same genetic code would be shared by mother, grandmother, great-grandmother, great-great-grandmother, and so on.” At all points in this chain, sons possess mitochondrial DNA received from their mothers, but sons cannot pass this mitochondrial DNA along to their daughters or sons. Thus, as a tool for establishing identity, mitochondrial DNA can be used to identify a woman anywhere in a vertical chain of women descended from one another. And it can identify a son of one of these women. But it cannot continue through the male line; with sons the chain is broken.
Gill and Ivanov extracted mitochondrial DNA from the nine bone samples brought from Russia. The extracts were amplified to workable quantities by using PCR. To their delight, the quality of the DNA sequences obtained, said Gill, was “generally comparable to that produced from fresh blood samples.” Focusing on two different stretches of the DNA sequence normally hypervariable between different humans, and deriving between 634 and 782 base pair letters for each of the nine subjects, the scientists achieved DNA profiles for all nine of the bone samples they possessed.
Next, they needed contemporary DNA to make comparisons. The search for living relatives began. People at the FSS and the Home Office drew books from libraries and pored over genealogical trees. Someone drew up a list of names of people who would be scientifically suitable and might be approached. In the case of the Empress Alexandra, finding a genetically useful living relative was easy. Alexandra’s older sister, Princess Victoria of Battenberg, had a daughter, who became Princess Alice of Greece. Princess Alice, in turn, produced four daughters and a son. In 1993, only one of these daughters, Princess Sophie of Hanover, was living. The son was Prince Philip, who became Duke of Edinburgh and the consort of Queen Elizabeth II of England. Prince Philip, Empress Alexandra’s grandnephew, was perfectly suited for a mitochondrial DNA comparison with bone material of the murdered Russian empress. Accordingly, Dr. Thompson, director of the FSS, wrote to Buckingham Palace and asked whether the prince would be willing to help. Philip agreed, and a test tube filled with his blood soon made its way to Aldermaston. The testing was done in those parts of the mitochondrial DNA sequence where the greatest variety between family groups occurs. By November, Gill and Ivanov had results: the match was perfect; the sequence of DNA base pairs between the mother, the three young women, and Prince Philip was identical. Gill and Ivanov knew that they had located the remains of Alexandra Feodorovna and three of her four daughters.
Confirming the presence of Tsar Nicholas II was far more difficult. The search for DNA material to compare with that extracted from the femur of Body No. 4 was widespread, prolonged, and, in several instances, controversial. The first attempts were made by Pavel Ivanov. It occurred to him that Nicholas II’s younger brother Grand Duke George, who died in 1899 of tuberculosis at the age of twenty-eight, was buried in the mausoleum of the Romanovs, the Cathedral of St. Peter and St. Paul in St. Petersburg. Comparison of DNA between brothers would nicely suffice. From England, Ivanov contacted Anatoly Sobchak, the mayor of St. Petersburg, and Vladimir Soloviev, who would become the investigator assigned to the Romanov case. “They protested that it would be too expensive,” Ivanov recalled. “ ‘The tombs in the fortress are made of Italian marble.… You must break it.… Who will pay for this?’ And so on.” For eight months, Ivanov persisted, and, at one point, Mstislav Rostropovich, the cellist and conductor, who is a friend of Sobchak, seemed about to pay for the exhumation of Grand Duke George.
Before this happened, however, Rostropovich told Ivanov that he was about to set out on a visit to Japan. Ivanov, still in England, remembered that in 1892 Nicholas II as tsarevich had visited Japan. In Otsu, the heir to the Russian throne suddenly had been attacked by a sword-wielding Japanese. The blow, aimed at his head, glanced off his forehead, bringing a gush of blood but failing to bite deep. The wound was bound with a handkerchief. For one hundred years, a museum in Otsu had kept the blood-soaked handkerchief in a small box. For DNA comparison purposes, nothing could provide more accurate positive identity than achieving a match between bone material of unknown origin and blood from a known person. Ivanov was eager to go to Japan, but, as always, “there was no money. The English said, Why should we pay for this? The Russians said, We have no money.” Eventually, Rostropovich arranged for Ivanov’s trip. “It was the money we were going to use to dig up George,” Ivanov said. “So, instead of George, we did Japan.”
The Japanese were not anxious to give up or even to disturb the handkerchief, but Rostropovich spoke to his friend the emperor of Japan, and the emperor spoke to the relevant authorities. When Ivanov arrived he was permitted to remove and take with him a strip of the handkerchief three inches long and one eighth of an inch wide. Unfortunately, back in Gill’s laboratory in England, Ivanov ran into difficulties. “The handkerchief had been handled by too many people,” he said. “Cells slough off from fingers. There was a lot of blood on the handkerchief, but who knows how much of it was Nicholas’s? And there was a lot of dust and dirt. It would be impossible to say that any result you got from that handkerchief was reliable. There were too many other possible contaminants.”
Having failed with both George and Japan, Ivanov came up with a third possible source of DNA for comparison to the piece of the presumed tsar’s femur at Aldermaston. In 1916, Nicholas II’s younger sister Grand Duchess Olga married Colonel Nicholas Kulikovsky, a commoner. With Kulikovsky, Olga had two sons, Tikhon, born in 1917, and Guri, born in 1919. In 1948, Olga and her family moved to Canada, where Kulikovsky bought a farm and raised cattle and pigs. Guri Kulikovsky died, but in 1992, when Gill and Ivanov began their work together, Tikhon, at seventy-five, was living in retirement in Toronto. He was, by that time, Tsar Nicholas II’s only living nephew and, as such, the best available source for comparative mitochondrial DNA. If the femur from Body No. 4 had belonged to Nicholas II, it should match perfectly with DNA from Tikhon Kulikovsky.
Mr. Kulikovsky, however, refused to cooperate. When Ivanov wrote to him, explaining the purposes of the investigation and asking for a blood sample, he received no reply. Ivanov tried again through Bishop Basil Rodzianko of the Orthodox Church in America, and, finally, through Metropolitan Vitaly, the head of the Russian Orthodox Church Abroad. Ultimately, Kulikovsky replied to Ivanov. “He told me he believed this whole bones business was a hoax,” Ivanov recalled. “He said, ‘How can you, a Russian man, be working in England, which was so cruel to the tsar and to the Russian monarchy?’ He said, ‘For political reasons, I will never give you a sample of my blood or hair or anything.’ ” Ivanov was disappointed, but he did not give up. “At that time, it was critical,” Ivanov said. “He was the closest relative. I spent a lot of my own money talking with him and his wife by telephone, assuring them that I was not a KGB agent. And they replied, ‘Then probably the only reason for your investigation is to prove that Tikhon Nicholaevich is not of royal blood.’ ” Ivanov gave up. “Okay, so we forgot about this Tikhon,” he said. “And after we published our work, some people wrote that our analysis was not accurate because we didn’t use the blood of Tikhon Kulikovsky. The fact is that his blood is no longer necessary. We found two other relatives. They gave us their blood, and we had everything we needed for our research.”
To locate the other two relatives, the Aldermaston genealogists looked again at the family tree. Because the chain of similar mitochondrial DNA is repeated indefinitely down through generations of females, they focused on the women closest by blood to Tsar Nicholas II. Beginning with his mother, Dowager Empress Marie, they found an unbroken line of five generations of mothers and daughters leading to a contemporary descendant willing to help. The tsar’s sister Grand Duchess Xenia had one daughter, Princess Irina. This Irina married Prince Felix Yussoupov, famous for having murdered Rasputin. Irina and Felix produced one child, a daughter, also named Irina. This second Irina married Count Nicholas Sheremetyev, with whom she had one child, a daughter, Xenia. Upon her marriage, young Countess Xenia Sheremetyeva became Xenia Sfiris. Now in her early fifties, Mrs. Sfiris lives in Athens and Paris, and it was in Athens that she received the FSS’s appeal for help. An exuberant, warmhearted woman, she agreed immediately. Following instructions, she pricked her finger, let some blood run into a paper handkerchief, where it dried, put the handkerchief in an envelope, and took it to the British Embassy. From there, it went, via diplomatic pouch, to Aldermaston.
The other donor of DNA material given to identify Nicholas II was found on what must seem an infinitely remote branch of the massive European royal family tree. Nevertheless, although the line stretched back over six generations, the connection was as reliable and productive as it was in the case of Mrs. Sfiris. James George Alexander Bannerman Carnegie, third Duke of Fife, Earl Macduff, and Lord Carnegie, is a sixty-six-year-old Scottish nobleman and farmer who descends from a common female ancestor of Tsar Nicholas II. She was Louise of Hesse-Cassel, a German princess who married King Christian IX of Denmark. One of her daughters became Empress Marie Feodorovna of Russia, the mother of Nicholas II. Another, older daughter, Alexandra, married the Prince of Wales, later King Edward VII. Queen Alexandra’s daughter Louise married the first Duke, of Fife. In 1929, Louise’s daughter Maud produced James, who, in 1959, succeeded to the title. The duke was willing to donate blood but, not wishing to incur publicity, made it a condition that he remain anonymous. Inevitably, in the course of time and with an investigation of this significance, knowledge leaked out.
As Gill and Ivanov expected, Xenia Sfiris’s mitochondrial DNA matched perfectly with that of the Duke of Fife. But when the matching 782 base pair letters lengths of the Greek woman and the Scottish peer were compared to the same section in the mitochondrial DNA extracted from the presumed tsar, there was a mismatch. A single letter was different. At a position numbered 16169, Xenia Sfiris and the Duke of Fife had a T; in this position Nicholas had a C. The other 781 pairs were in identical sequence. To check their data, Gill and Ivanov did a second mitochondrial DNA extraction from the bone believed to be the tsar’s. They cloned the DNA in this region after amplifying it with PCR and then transformed the product into E. coli bacteria. When fresh sequencing of these new clones was performed, seven of the clones were found to have a T at position 16169, thus matching Mrs. Sfiris and the Duke of Fife. But twenty-eight clones still presented the single spelling mistake, a mismatching C. The Aldermaston scientists concluded that Tsar Nicholas II had possessed two forms of mitochondrial DNA, one of which matched his relatives exactly; the other, at a single point, did not. This rare condition is known as heteroplasmy.
This single mismatched base pair letter caused great anxiety at Aldermaston. In their paper, the two scientists presented their interpretation of what they had found: “We consider … that the mitochondrial DNA extracted from the tsar was genetically heteroplasmic. This complicates the interpretation because the strength of the evidence depends upon whether we accept a priori that a mutation has occurred in the tsar. The probability of a single mutation was calculated to be approximately one in three hundred per generation, but this estimate does not take account of the incidence of heteroplasmy (much of which may be undetected).”
Gill understood that this one-letter mismatch raised questions about the validity of his findings. He believed that a mutation did occur, although he admitted that the odds against a mutation in any given generation were long. “A mutation is thought to occur [in a family] about once in three hundred generations,” he said. But he insisted that he is talking primarily about a heteroplasmy, which he found, not a mutation, which he could not prove but which is the probable cause of that heteroplasmy. “Heteroplasmy is different from a mutation in nuclear DNA; it means that there are two types of mitochondrial DNA in the same person. What we did was demonstrate that there are two types of mitochondrial DNA in the tsar. One of these types differed by just one base; the other was identical to the relatives.’ That’s pretty good evidence that there has been a real mutation. Bear in mind that we’re working right at the frontiers of knowledge. The actual incidence of this type of phenomenon is not really known, and we suspect that it is much more common than originally had been anticipated.”
In July 1993, after ten months of work, Gill and Ivanov were ready to announce their results to the world. The Forensic Science Service convened a press conference and, on July 10, a large hall at the bleakly modern Home Office building in Queen Anne’s Gate was filled with reporters, photographers, and television cameramen. Dr. Janet Thompson, the director general of the FSS, presided. Aware that questions might be asked as to who had borne the cost of this research, she began by expressing hope that “the FSS will soon be able to put the techniques used, once validated, into practice in criminal casework to the benefit of the criminal justice system as a whole.”
Gill explained what he and his colleagues had done. He described how the sex of the specimens had been determined, how the family relationship between five skeletons had been established, how Prince Philip’s blood had made certain the identity of Alexandra Feodorovna and her daughters, and how the heteroplasmy found in the tsar’s DNA had complicated the effort to make an absolute statement about Nicholas. Nevertheless, the Aldermaston team announced that, given the DNA evidence and adding it to the anthropological and historical evidence provided by others, they were 98.5 percent certain that these were the Romanovs. This percentage, Gill said, was based on the most conservative interpretation of the DNA evidence. A more generous interpretation would increase the probability to 99 percent. Pavel Ivanov took a broader view of what had been done. “We are very close to the last part of this mystery, to one of the great mysteries of the twentieth century, one of the great mysteries of my country, of Russia,” he said.
The press conference produced headlines: RIDDLE OF ROMANOV REMAINS IS SOLVED (Financial Times), DNA TESTS IDENTIFY TSAR’S SKELETON (The Times), TSAR NICHOLAS’S BONES IDENTIFIED (The Washington Post). Tass told Russian readers that “British scientists” were “almost without any doubt” that the remains found in Siberia were those of Tsar Nicholas II and his family. Seven months later, in February 1994, Peter Gill, Pavel Ivanov, and others put their findings in print in their own words, publishing a description of their work in Nature Genetics, the authoritative journal of their profession. Their findings and article have never been challenged or even mildly criticized, in print or orally, by another DNA scientist.
* McCrery, a florid, enthusiastic man who was a policeman before he went to Cambridge to study Russian history, takes considerable credit for engineering the British-Russian effort. Hearing about the discovery of the bones, he says, he rang up Avdonin in Ekaterinburg. Avdonin put him on to Pavel Ivanov. Ivanov told him that the best place in the world for DNA testing was Aldermaston and gave him the name of Peter Gill. McCrery telephoned Gill, who, he says, was “quite excited, but not sure the Home Office would approve. Well, Kenneth Clark, [then] the home secretary, lives around the corner from me and I’ve known him for years. He is my M.P. So I contacted him and pointed out how prestigious it would be for the Forensic Science Service to be involved. ‘Will you give permission?’ And Clark said, ‘It’s a wonderful idea.’ So then I rang Ivanov back and Ivanov said, ‘How do I get there? I don’t have any money.’ So I said, ‘I’ll pay for it.’ Actually, somebody in Russia paid for Ivanov’s trip, but I got in touch with Applied Biosystems, which makes gene scanners and other machines they use in DNA work, and asked them whether they would pay Ivanov’s expenses in England. They said yes, and they came up with three to five thousand pounds for him to live on for ten months. So he came and brought the bones.”
* In fact, the security men of both monarchs were so concerned about the possibility of terrorism that King Edward never actually set foot on the soil of the Russian Empire. All meetings were held on the two yachts.