Mitochondrial DNA

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Mitochondrial DNA Y-chromosome markers
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Mitochondrial DNA

• - Multiple organelles in the cytoplasm of the
  cell
• - Powerhouses of the cell - oxidative
  respiration of the cell ? use oxygen provide
  energy fuels multiple reactions.
• - Found outside the nucleus
• Own DNA ? different than genomic DNA
• Smaller
• Single circular, double-stranded molecule
  16,569 bp
• Contains 37 genes 13 protein-coding, 3
  ribosomal RNA and 22 transfer RNA genes
• Only a small region is non-coding
• Inherited from mother ? all mitochondria from
  egg mtDNA type cannot be heterozygous, or
  exhibit two different types no contribution
  from the father

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• Identity marker for maternal relatives
• D-loop mutate with high frequency useful for
  forensic
• 1-2 variation between unrelated individuals
  one to two nucleotides out of every hundred is
  different highly variable between unrelated
  individuals
• High mutation rate single individual show a
  mixture of types - heteroplasmy
• Multiple copies more copies of mtDNA than
  nuclear DNA
• Useful in badly degraded sample insufficient
  nuclear DNA mtDNA last typable DNA left

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• Advantages
• Extremely small amounts of DNA material needed
• Degrading of DNA slower
• Useful in tracing family lineages

• Limitations
• Cannot be used to distinguish between sibship or
  maternal relatives
• Discrimination power ? limited to size of
  database Anderson sequence
• Heteroplasmy can complicate analysis

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Example Tsar Nicholas II

• Nicholas II assumed the throne 1894
• Married to Alexandra (german)
• Nicholas was influenced by Alexandra who was
  influence by Rasputin (monk) and Kaiser Wilhelm.
  Who ruled Russia?
• Four daughters Olga ,Tatiana, Maria Anastasia
  One son Alexei, born 12 Aug 1904 (hemophilia)
• 1 Aug 1914 war with Germany. Nicholas role of
  commander-in-chief (September 1915)
• March 1917 February Revolution ? abdication
  of Nicholas II

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• Provisional Russian Government kept Nicholas
  and family The Alexander palace
• Kerensky government - Tobolsk in Siberia (August
  1917)
• Bolshevik Revolution November 1917 moved to
  Red-controlled Yekaterinburg.
• 16 / 17 July 1918 Family, their physician and
  three servants were executed in the basement of
  the house Ipatiev, by Yakov Yurovsky (Bolshevik)

• Bodies disposed in mineshaft Four Brothers
• Yurovsky hearing rumours moved bodies to
  another mineshaft
• Vehicle broke bodies buried in a sealed and
  consealed pit on Koptyaki road, 12 miles north
  of Ekaterinburg (except for two bodies)
• Claims from various people one of Romanov
  children (Anna Anderson Anastasia)

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• Report by Yurovsky came to light in 1970s,
  public knowledge 1990s
• July 1991 President Boris Yeltsin authorized
  the exhumation
• Almost 1000 bones five female and four male
  skeletons were found ? two were missing
• William Maples dental and bone specimens ?
  Alexei and Anastasia were missing
• Pavel Ivanov and Peter Gill nuclear DNA
  fingerprinting and mtDNA test

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DNA fingerprinting test results

• Five of the bodies related (3 siblings)

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mtDNA test results

• Reference sample for Alexandra Prince Philip
  (grandnephew of Alexandra) submitted sample.
• The mtDNA match for the 3 daughters, Alexandra
  and Prince Phillip

• Nicholas mtDNA rare anamoly (heteroplasmy)
  one base position two different bases were found
  (C and T at one position).
• Countess Xenia Cheremetiff-Sfiri
  (great-granddaughter of the Csars sister ) and
  the Duke of Fife showed only a T
• Grand Duke Georgij (died of tuberculosis in
  1899) brother of Tsar Nicholas II, were exhumed
  in July 1994
• Heteroplasmy in both rare mixture of C and T
  at position 16169
• Anna Anderson not Anastasia
• According to Yurovsky Alexei and Anastasia
  were burnt near the burial site and their ashes
  scattered and concealed.

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Y Chromosome markers

• Nuclear DNA ? paternal inheritance (father to
  son)
• Valuable as lineage markers family
  relationships.
• Useful in analysis of semen stains - mix samples

• Advantages
• Useful in tracing family relationships among
  males
• Lack of recombination, - markers useful to
  measure relatedness of individuals with a common
  geographical origin.

• Limitations
• Discriminatory power limited by the size of the
  database

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Example

• Swinoujscie Poland ? 14 girls raped and
  murdered
• Information of offender scanty
• Special police group of 8 established
• Men between 22 38 years in Swinoujscie area ?
  investigated
• 714 suspects interrogated ? 421 DNA typed
• 28-year old man Y-chromosome markers fitted
  (autosomal STRs excluded)
• Brother of man arrested ? autosomal STRs matched.

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Results
Y-chromosome DNA profiles in samples from the
five cases
DYS locus
Samples analysed
DYS 19
DYS 393
DYS 390
DYS 392
DYS 391
DYS 389II
DYS 389I
DYS 385I / II
Case 1
11
16
13
10
13
29
11 / 15
25
29
11 / 15
Case 2
11
16
13
10
13
25
Case 3
13
16
10
13
29
25
11 / 15
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Case 4
11
15 / 16
12 / 13
10
29 / 30
24 / 25
12 / 13
11/14/ 15/17
11
10
13
29
11 / 15
Case 5
16
13
25
Dominating mixture in the result
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Results
Deduced autosomal profile of the rapist
(ProfilerPlus) in three of the cases
ProfilerPlus loci
Analysed samples
D13S317
D3S1358
V W A
FGA
D5S818
AMG
D8S1179
D21S11
D18S51
D7S820
29/30/31
15/18/20
Case 1
15/16/17
14/15/17
22 / 22.2 / 23
11/12/13
10/11/12
8/10/12
XgtY
12 / 13
Case 2
15/16/17
14/17
21 / 22 / 22.2 / 25
10/11/12
10/11/14
8 / 9 / 10 / 12
XgtY
12/13/14
28 / 29 / 31 / 31.2
17 / 20
Case 3
28/29/31
14/17/20
15/16/17
14/17/ 18
20 / 22 / 22.2 / 23
11/12
9 / 10 / 11 / 12
7 / 8 / 12 / 13
XgtY
12 / 13
Deduced Rapists profile
14 / 17
16 / 17
22 / 22.2
11 / 12
10 / 11
8 / 12
XY
12? / 13
29 / 31
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Alleles not present in the victims reference
material
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Results
Comparison of rapists DNA profile (ProfilerPlus)
with sample originating from suspect and the one
of his brother with an identical Y-haplotype
ProfilerPlus loci
Samples analysed
D13S317
D3S1358
V W A
FGA
D5S818
AMG
D8S1179
D21S11
D18S51
D7S820
Suspect (KW)
15 / 20
16 / 17
10 / 12
10 / 11
XY
12 / 13
30.2 / 31
17
22
10 / 11
Suspects brother (TW)
14 / 17
16 / 17
22 / 22.2
11 / 12
10 / 11
8 / 12
XY
12 / 13
29 / 31
20
Deduced Rapists profile
14 / 17
16 / 17
22 / 22.2
11 / 12
10 / 11
8 / 12
XY
12? / 13
29 / 31
20
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Comparison of DNA typing methods
High
RFLP Single and multi-locus probes
Multiplex STRs
Power of discrimination
(Genetics)
ABO blood groups Polymarker D1S80 Single STR DQa
mtDNA
Low
Speed of analysis
Fast
Slow
(Technology)
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The End