presentation - final draft
TRANSCRIPT
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“Great” Grandma and You: Methods of Analyzing Human MtDNA Substitution Rate BY SETH NELSONTHURSDAY, OCTOBER 8TH, 2015
2Outline
I. Mitochondria and DNAII. MtDNA anatomy and replicationIII. Methods of finding substitution rateIV. Improvement on current findingsV. Using the rate
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I. Mitochondria and DNA
I. MITOCHONDRIA AND DNA
II. MTDNA ANATOMYIII. METHODS OF
FINDING SUBSTITUTION RATE
IV. IMPROVEMENT ON CURRENT FINDINGS
V. USING THE RATE
Mitochondria
Endosymbionts with proto-eukaryotes (Andersson et al., 2003)
Applications in forensics and evolutionary relatedness Need to know
mutation rates for accurate judgment
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Figures: Sadava et al. (2005)
5Using DNA for phylogenetic inference
Figure: García et al. (2011)
6Why use mtDNA
More mtDNA copies than nDNA (Robin and Wong, 1988)
Mitochondria are inherited from mother (Schwartz and Vissing, 2003)
High mutation rate, good for closely related individuals (Butler and Levin, 1998)
Image from https://www.thermofisher.com/us/en/home/technical-resources/research-tools/image-gallery/image-gallery-detail.2643.html
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II. MtDNA Anatomy and Replication
I. MITOCHONDRIA AND DNA
II. MTDNA ANATOMY AND REPLICATION
III. METHODS OF FINDING SUBSTITUTION RATE
IV. IMPROVEMENT ON CURRENT FINDINGS
V. USING THE RATE
8Anatomy of mtDNA
Figure: Pakendorf and Stoneking (2005)
Transfer RNAs
NADH Dehydrogenase subunits
Cytochrome c Oxidase subunits
Cytochrome bRibosomal RNAs
ATP Synthase subunits
9Control region
Controls replication
No protein product Two hypervariable
regions
Figure: Pakendorf and Stoneking (2005)
10Beginning of replication
Initiates at heavy strand origin
Light strand synthesis follows
Figure: Clayton (2000)
11Mutations in replication of DNA
Insertion Deletion Frameshift Substitution
Transition Transversion
Figure: Sadava et al., 2011
12Substitutions happen at specific rates
Substitutions per site per million years Numerator: Number of sequence differences,
only counting substitutions That is, no insertions, deletions, etc.
Denominator: Time since last common ancestor between sequences of comparison
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III. Methods of Finding Substitution Rate
I. MITOCHONDRIA AND DNAII. MTDNA ANATOMYIII. METHODS OF FINDING
SUBSTITUTION RATEI. ConsiderationsII. PedigreeIII. Phylogenetic
IV. IMPROVEMENT ON CURRENT FINDINGS
V. USING THE RATE
14Secondary structure forms
Light strand forms loop structure (Pereira et al., 2008)
Selective pressure on control region
Figure: Pereira et al. (2008)
15MtDNA can recombine
Mitochondria possess recombinase activity (Thyagarajan et al., 1996)
Does not affect substitution rate (Kraytsberg et al., 2004)
Figure: Thyagarajan et al. (1996)
16Some paternal inheritance
Single case of paternal inheritance in man (Kraytsberg et al., 2004)
Figure: Kraytsberg et al. (2004)
17Pedigree analysis is direct observation
Analyze mtDNA from closely related individuals English family with
Leber’s hereditary optic neuropathy
Age of last common ancestor is known with certainty
Figure: Howell et al. (2003)
18Less time means fewer mutations
Pedigree analysis tends to count fast mutations Potentially overestimate substitution rate
19Phylogenetic analysis uses equations
Analyze mtDNA from distantly related individuals Primates, back to
chimp and human CA
Figure: Hasegawa et al. (1993)
20Equations as estimates
Use of equations for rate Transition rate: Transversion rate: Substitution rate:
21More time means more uncertainty
Denominator more uncertain Phylogenetic analysis counts all substitutions
since last CA Reversions will cause undercount in mutations
Need methods of calibration
22Nodes vs. tips
Figure: Rieux et al. (2014)
23Calibration affects rates
Figure: Rieux et al. (2014)
24Noncoding region is higher than coding region
Pedigree rate is higher by order of magnitude Rates are in substitutions per site per million
years
Method
Noncoding
RegionCoding Region
Pedigree (99.5% CI)0.475 (0.265-
0.785)a0.15 (0.02-0.49)a
Phylogenetic (±1 Std
Error)
0.033 (0.027-
0.039)b0.0170 (--)c
Pedigree rates from Howell et al. (2003)Phylogenetic noncoding from Hasegawa et al. (1993)Phylogenetic coding from Ingman et al. (2000)
25Pedigree is higher than phylogenetic
Method Weighted ratePedigree 0.17Phylogenetic, Tip 0.021Phylogenetic, Node 0.018
Pedigree rate from Howell et al. (2003)Phylogenetic, tip-calibrated rate from Rieux et al. (2014)Phylogenetic, node-calibrated rate from Hasegawa et al. (1993) & Ingman et al. (2000)
*Rates are in substitutions per site per million years
26Context is everything (Pääbo, 1996)
Phylogenetic rate: Common ancestor is >100,000 years ago
Pedigree rate: Common ancestor in <10,000 years ago
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IV. Improvement on Current Findings
I. MITOCHONDRIA AND DNA
II. MTDNA ANATOMY AND REPLICATION
III. METHODS OF FINDING SUBSTITUTION RATE
IV. IMPROVEMENT ON CURRENT FINDINGS
V. USING THE RATE
28Bringing the rates together
Figure: Ho et al. (2005)
29Bringing the rates together
Figure: Ho et al. (2005)
30A better outgroup is in the nucleus
MtDNA integrated into nucleus
540 bp segment Identical in all
tested genomes (Zischler et al., 1995)
Figure: Zischler et al. (1995)
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V. Using the Rate
I. MITOCHONDRIA AND DNA
II. MTDNA ANATOMY AND REPLICATION
III. METHODS OF FINDING SUBSTITUTION RATE
IV. IMPROVEMENT ON CURRENT FINDINGS
V. USING THE RATE
32Use in forensics
Forensic applications focus on HV1 and HV2 Romanov identification (Butler and Levin,
1998) Tsarina, her daughters, Prince Philip were exact
matches One mismatch for Tsar Nicholas II and relatives “Anastasia” did not match
33Dating divergence
Estimating common ancestor of Neanderthals and Humans
Figure: Ho et al. (2005)
34Unrelated to “Great” Grandma
How far back in time do we need to go to be “unrelated” to our ancestors?
1.1% (12 bp) difference in unrelated control sequences (Piercy et al., 1993)
Roughly 1000 generations before we are unrelated to our ancestors
35Knowing this, we look deeper
Substitution rate is effectively variable Temporally and spacially
Allows a second look at archeological dates Could help us understand relationships
better Methods used in mtDNA could be extended
36Acknowledgements
Thank you! Friends Family Chris Cole and rest of biology faculty Everyone else here
37Questions?
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