Gene Cloning and DNA Analysis: An Introduction, Sixth Edition ...

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Figure 16.5
Part III The Applications of Gene Cloning and DNA Analysis in Biotechnology
Family tree showing the matrilineal relationship between
Prince Philip, Duke of Edinburgh and Princess Victoria
of Hesse, the Tsarina’s sister. Males are shown as blue
boxes and females as red circles.
Grand Duke
Ludwig
Princess Alice
Tsarina Princess Victoria
of Hesse
Alice of
Battenburg
Prince Philip,
Duke of Edinburgh
other unfortunate group of people? To address this problem, the DNA from the bones
was compared with DNA samples from living relatives of the Romanovs. This work
included studies of mitochondrial DNA, the small 16 kb circles of DNA contained in
the energy-generating mitochondria of cells. Mitochondrial DNA contains polymorphisms
that can be used to infer relationships between individuals, but the degree of
variability is not as great as displayed by STRs, so mitochondrial DNA is rarely used for
kinship studies among closely related individuals such as those of a single family group.
But mitochondrial DNA has the important property of being inherited solely through
the female line, the father’s mitochondrial DNA being lost during fertilization and not
contributing to the son or daughter’s DNA content. This maternal inheritance pattern
makes it easier to distinguish relationships when the individuals being compared are
more distantly related, as was the case with the living relatives of the Romanovs.
Comparisons were therefore made between mitochondrial DNA sequences obtained
from the skeletons and that of Prince Philip, the Duke of Edinburgh, whose maternal grandmother
was Princess Victoria of Hesse, the Tsarina Alexandra’s sister (Figure 16.5).
The mitochondrial DNA sequences from four of the female skeletons—the three
children and the adult female identified as the Tsarina—were exactly the same as that
of Prince Philip, strong evidence that the four females were members of the same lineage.
Comparisons were also made with of two living matrilineal descendants of Tsar
Nicholas’s grandmother, Louise of Hesse-Cassel. This analysis was more complicated,
as two sequences were present among the clones of the PCR product obtained from the
adult male thought to be the Tsar. These sequences differed at a single position which
was either a C or a T, the former four times more frequent than the latter. This could
indicate that the sample was contaminated with somebody else’s DNA but instead was
interpreted as showing that the Tsar’s mitochondrial DNA was heteroplasmic, an infrequent
situation where two different mitochondrial DNAs co-exist within the same cells.
The two descendents of the Tsar’s grandmother both had the mitochondrial DNA
version with a T at this position, suggesting that the mutation producing the C variant
had occurred very recently in the Tsar’s lineage. Support for this hypothesis was subsequently
provided by analysis of DNA from the Tsar’s brother, Grand Duke George
Alexandrovich, who died in 1899, which showed that he also displayed heteroplasmy
at the same position in his mitochondrial DNA. On balance, the evidence suggested that
the Tsar’s remains had been correctly identified.