mobile element applications for molecular ecology
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Mobile Element Applications for Molecular Ecology. David A. Ray West Virginia University. Outline. An introduction to mobile elements Classification, distribution, evolution, impacts SINEs as phylogenetic tools Primate phylogeny SINEs as tools in population genetics and forensics - PowerPoint PPT PresentationTRANSCRIPT
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Mobile Element Applications for Molecular Ecology
David A. Ray
West Virginia University
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Outline
An introduction to mobile elements Classification, distribution, evolution, impacts
SINEs as phylogenetic tools Primate phylogeny
SINEs as tools in population genetics and forensics Population-indicative insertions and narrowing
a criminal investigation Current work
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Outline
An introduction to mobile elements Classification, distribution, evolution, impacts
SINEs as phylogenetic tools Primate phylogeny
SINEs as tools in population genetics and forensics Population-indicative insertions and narrowing
a criminal investigation Where to go from here?
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• Present in most eukaryotes• Very high copy numbers
• 10,000 – 1 million+ copies/genome • Retrotransposons
• SINEs (Short INterspersed Elements )• LINEs (Long INterspersed Elements)
• Transposons and derived elements• hAT (hobo, Activator, Tam) elements• MITEs (Miniature Inverted Terminal-
repeat Elements)
An Introduction to Mobile Elements
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1. Short INterspersed Elements2. <500 bp3. No open reading frame (non-coding)4. Copy-and-paste mobilization5. Derived from tRNA or 7SL RNA6. Alu (primates), B1 (rodents), SmaI
(salmon), AFC (cichlids), etc…7. Very high copy numbers
(>100,000 copies/genome)
What are SINEs?
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SINE Mobilization – “Copy and Paste”
Pol III transcription
Reverse transcription and insertion
1. Usually a single or a few ‘master’ elements
2. Pol III transcription to an RNA intermediate
3. Target primed reverse transcription (TPRT) – enzymatic machinery provided by LINEs
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Genomic Impacts:SINE Insertions and Mutation
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Genomic Impacts:SINE/SINE Recombination
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Mammalian SINE Distribution
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Genome
Mobile element subfamily evolutionT
ime
Subfamily 1
Subfamily 2
Subfamily 3G
enom
e
Organismal speciation event
Gen
ome
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SINE Subfamilies – Diagnostic Mutations
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SINE Subfamilies – Diagnostic Mutations
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Outline
An introduction to mobile elements Classification, distribution, evolution, impacts
SINEs as genetic markers Primate phylogeny
SINEs as tools in population genetics and forensics Population-indicative insertions and narrowing
a criminal investigation Where to go from here?
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1. Identical by descent2. Known ancestral state 3. Simple evolutionary model4. Neutral5. “Low-tech”6. Bi-allelic markers7. Essentially homoplasy-free8. “Two markers in one”
Presence/absence andDNA sequence
Mobile elements as tools for conservation biologists
Alu Insertion PCR AssayAlu Insertion PCR Assay
400400 bpbp
100100 bpbp
332211
400400 bpbp BandBand 100100 bpbp BandBand400 & 100400 & 100 bpbp BandsBands
AA
BB
Ray et al., Forensic Sci. Intl. 2005
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Identity by Descent
Species A
Species A
Species A’ Species B
Identity By Descent
time
Identity By State
Species AATGGTCC
Species BATGATCC
Species AATGGTCC
Species BATGGTCC
insertion
mutation
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SINE analysis of phylogeny
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Misleading and homoplasy-inducing events in a SINE analysis.
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Rates of misleading events in SINE analysis
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Outline
An introduction to SINEs SINEs as genetic markers
Primate phylogeny SINEs as tools in population genetics and
forensics Population-indicative insertions and narrowing
a criminal investigation Where to go from here?
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Platyrrhine Primates
16 genera described as six “natural groups”
Callitrichids
Capuchins and squirrel monkeys
Owl monkeys Saki monkeysAtelids Titi monkeys
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Platyrrhine Primates
16 genera described as six “natural groups”
Callitrichids
Capuchins and squirrel monkeys
Owl monkeys Saki monkeysAtelids Titi monkeys
Family Cebidae:Callithrix, Cebuella, Leontopithecus, Saguinus, Callimico, Cebus, Saimir, Aotus
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Platyrrhine Primates
16 genera described as six “natural groups”
Callitrichids Atelids
Capuchins and squirrel monkeys
Owl monkeys Saki monkeysTiti monkeys
Family Atelidae:
Ateles, Lagothrix, Brachyteles, Alouatta
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Platyrrhine Primates
16 genera described as six “natural groups”
Callitrichids Saki monkeys
Capuchins and squirrel monkeys
Owl monkeys Atelids Titi monkeys
Family Pitheciidae:
Pithecia, Chiropotes, Cacajao, Callicebus
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Cebidae
Pitheciidae
Atelidae
Previous Molecular Analyses
G6PD - Parsimony G6PD - Likelihood G6PD - Bayesian
E-globin - ParsimonyIRBP - Parsimony
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Investigating Platyrrhine Phylogenetics with Alu Elements
•Two complementary methodologies:
Experimental – Use “wet-bench” techniques to find Alu elements in nine New World monkey genomes
Advantages – Nine whole genomes to search, no chromosomal limitations
Disadvantages – Nine whole genomes to search, cost, selection bias
Computational – Use available sequence databases to identify and characterize Alu elementsAdvantages – Quick and inexpensive, no biasDisadvantages- Limited to available sequence data and
organisms (three)
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Alu Recovery – Experimental (9 taxa)
Mol. Phyl. Evol. (In press)
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Alu Recovery – Computational (3 taxa)(NISC comparative vertebrate sequencing
program; http://www.nisc.nih.gov/)
Query sequence - Callithrix Human ortholog
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Results – Alu Loci
Alu insertions were considered “usable” if they amplified as discreet bands in 5 of the 9 available platyrrhine genera and at least one Catarrhine primate.
Experimental – 89 usable loci from nine taxaCallithrix, Saguinus, Saimiri, Aotus,
Pithecia, Callicebus, Ateles, Alouatta, Lagothrix
Computational – 94 usable loci from three taxaCallithrix, Pithecia, Saimiri
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Results – Insertion Polymorphisms
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New World Monkey Phylogeny Resolved with 185 Alu Insertion Polymorphisms
Consistency index = 1.00Homoplasy index = 0.00
Mol. Phyl. Evol. (2005)G6PD - Parsimony G6PD - Likelihood G6PD - Bayesian
E-globin - ParsimonyIRBP - Parsimony
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Finding mobile element loci for a group
Mobile element subfamilies tend to have finite life spans
The first step in identifying polymorphic and/or species/taxon specific markers is to identify the appropriate subfamily
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Alu evolution in New World primates Bayesian analysis of 60 Alu sequences
found exclusively in platyrrhine primate genomes
Well-supported clades were analyzed for diagnostic sites
Ray and Batzer, BMC Evol Biol – 2005
Alu evolution in New World Primates
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Three (possibly five) new platyrrhine-specific subfamilies were characterized – AluTa5, AluTa10, AluTa15 Two subfamilies are exclusive to Cebid and Atelid monkeys In addition, we identified a unique mode of Alu subfamily evolution
Ray and Batzer - BMC Evol Biol (2005)
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AluSp-like
AluSc-like
A unique mode of Alu subfamily evolution
Ray and Batzer - BMC Evol Biol (2005)
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A unique mode of Alu subfamily evolutionT
ime
AluSc
AluT
AluTa10, AluTa15
AluSp
Ancestral primate genome
Ancestral platyrrhine genome
Cebid-Atelid genomes
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A
B
C. k
uh
lii
C. p
ygm
aeu
s
Sag
uin
us
Sai
mir
i
Ao
tus
wat
er
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0.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
Platyrrhini (n=58) Cebidae-Atelidae (n=14)
Cebidae (n=6) Callitrichinae(n=40)
Callithrix (n=40)
Sc
Sg
Sp
Sq
Sx
Ta10
Ta15
Ta7
Y
A
B
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Outline
An introduction to SINEs SINEs as genetic markers
Primate phylogeny SINEs as tools in population genetics and
forensics Population-indicative insertions and narrowing
a criminal investigation Where to go from here?
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SINEs, Biology, and Investigative Forensics
1) Trace DNA detection 2) Species identification3) Human DNA quantitation4) Human gender typing5) Inferring geographic affiliation
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Human Population Biology and Investigative Forensics
1) Trace DNA detection 2) Species identification3) Human DNA quantitation4) Human gender typing5) Inferring geographic affiliation
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FBI CODIS loci are great for matching a single unknown DNA sample to a single individual.
Must have a suspect. Early in an investigation, can we narrow the
field of suspects using information from Alu polymorphisms?
Human Population Biology and Investigative Forensics
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Human Population Biology and Investigative Forensics
Mobile elements continue to propogate in the human genome.
Many elements are polymorphic and occur at variable frequencies in human populations.
Display-based PCR methods can be used to “extract” recent, population-indicative elements.
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Inferring Geographic Affiliation
1) Series of genetic markers (100 Alu loci)
2) Database of human variation (currently 715 individuals of known ancestry)
3) Genotype unknown sample(s) 18 samples from two forensic laboratories
4) Analytical approach (Structure analysis)Forensic Sci. Intl. (In press)
Forensic Sci. Intl. (In press)
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Identifying 18 Unknown DNAs
Forensic Sci. Intl. (In press)
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Outline
An introduction to SINEs SINEs as genetic markers
Primate phylogeny SINEs as tools in population genetics and
forensics Population-indicative insertions and narrowing
a criminal investigation Where to go from here?
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Where Do We Go from Here?Population Biology and Phylogenetics
Expand the use of mobile elements as phylogenetic and population genetic markers to additional organisms Genome exploration Mobile element characterization Techniques for identifying “useful” loci Species identification and DNA quantitation Bats, blowflies, fish, and crocodilians
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Contributors
Batzer Lab - LSUBatzer Lab - LSUMark A. Batzer Scott Herke Jinchuan Xing Michael A. HallDale J. Hedges Meredith E. LabordeJerilyn Walker Bridget A. AndersAbdel-Halim Salem Brittany R. WhiteRandy Garber Nadica StoilovaRichard Cordaux Justin D. FowlkesGail Kilroy Cheney Huang
Jorde Lab – University of UtahJorde Lab – University of UtahLynn Jorde David Witherspoon
Center for Reproduction of Endangered Species – San Diego ZooCenter for Reproduction of Endangered Species – San Diego Zoo
Oliver A. Ryder Leona G. Chemnick
Texas Tech UniversityTexas Tech UniversityLou Densmore
University of FloridaUniversity of FloridaGinger Clark
Savannah River Ecology LaboratorySavannah River Ecology LaboratoryTravis Glenn
WVUWVUJeffrey Wells, Zena Urban,
Heather Campbell
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Future Directions
Additional Loci: Tiered approach to subpopulation affiliations.
Initial screen for broad population affiliation Secondary screen for subpopulation affiliation
Loci good for the first screen unlikely to be useful in the second.
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Allele freq = 0.9
Allele freq = 0.2Allele freq = 0.3
Allele freq = 0.3
Population-indicative Loci
Little variation to exploit at subpopulation levels
Subpopulation-indicative loci are needed