genetic coalescence in north american white pines kathleen farrell dr. aaron liston, dr. richard...
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Genetic Coalescence in North American
White PinesKathleen Farrell
Dr. Aaron Liston, Dr. Richard Cronn,
John Syring
P. strobus
Coalescence is the pattern of common ancestry.
Coalescence theory: At some point all alleles in the population will be traceable to a single common ancestral allele that existed in the population at time t0.
Factors: Gene flowHybridizationSelection Population size
Objective: to find low copy nuclear genes that accurately demonstrate the relationships (phylogeny) between closely related species, North American White Pines.
AA
BB
CC
Coalescing Tree:
AAC
BB
C
Non-Coalescing Tree:Why low copy genes? Orthologs vs. Paralogs
Objective: to find low copy nuclear genes that accurately demonstrate the relationships (phylogeny) between closely related species, North American White Pines.
AA
BB
CC
Coalescing Tree:
AAC
BB
C
Non-Coalescing Tree:Why low copy genes? Orthologs vs. Paralogs
Species Sampled:
North American White PinesP. albicaulis Whitebark PineP. ayacahuite Mexican White PineP. chiapensis Chiapas White PineP. flexilis Limber PineP. lambertiana Sugar PineP. monticola Western White PineP. strobiformis Southwestern White PineP. strobus Eastern White Pine
OutgroupsP. monophylla Singleleaf PinyonP. longaevaBristlecone Pine
P. longaevaP. monophylla
P. flexilis
P. ayacahuite P. albicaulis
P. lambertiana
DNA Extraction from the megagametophyte tissue.
megagametophyte (1n)
embryo (2n) shoot end
embryo root tip (radicle)
Methods and Approach:
How it works:
DNA DenaturesPrimers AnnealTaq Polymerase builds
a complementary copy.
Polymerase Chain Reaction (PCR)
How it works:
DNA DenaturesPrimers AnnealTaq Polymerase builds
a complementary copy.
Polymerase Chain Reaction (PCR)
Polymerase Chain Reaction (PCR)
Results: Gel Electrophoresis
IFG8612 Optimization: MONTICOLA 02S1
Temperature (C) 56 57 58 59 60 61
Temperature (C) 56 57 58 59 60 61
62 63 64 65 66 67 68 62 63 64 65 66 67 68
Contr
ol
Contr
ol
2.0 mM MgCl 1.5 mM MgCl
Polymerase Chain Reaction (PCR)
Results: Gel Electrophoresis
IFG8612 Optimization: MONTICOLA 02S1
Temperature (C) 56 57 58 59 60 61
Temperature (C) 56 57 58 59 60 61
62 63 64 65 66 67 68 62 63 64 65 66 67 68
Contr
ol
Contr
ol
2.0 mM MgCl 1.5 mM MgCl
Polymerase Chain Reaction (PCR)
Results: Gel Electrophoresis
IFG8612 Optimization: MONTICOLA 02S1
Temperature (C) 56 57 58 59 60 61
Temperature (C) 56 57 58 59 60 61
62 63 64 65 66 67 68 62 63 64 65 66 67 68
Contr
ol
Contr
ol
2.0 mM MgCl 1.5 mM MgCl
Cycle SequencingAnalyzing Results:Clean Sequence
Sequence Noise
------------------------GTAATT----AGATTTGAATGGCATGGAATGAATGGAAA-GCaGGAtaTTGTCATTTCTTTCAGTAATT----AGATTTGAATGGCATGGAATGAATGGAAA----GGAtaTTATCATTTTTTTCaGTAATTAATTAGATTTGAATGGCATGGAATGAATGGAAA-GCaGGAtATTGTCATTTCTTTCAGTAATT----AGATTTGAATGGCATGGAATGAATGGAAA
Alignment and Editing
Developing a Phylogenetic Tree
Export alignments to PAUP* Search for phylogenetic trees based on maximum
parsimony using the branch and bound algorithm. Calculate bootstrap values (measure of confidence for
each branch) and branch length (measure of relative divergence).
Results:
Cinnamyl Alcohol Dehydrogenase
IFG8898IFG8612
Results:
Cinnamyl Alcohol Dehydrogenase
IFG8898IFG8612
Results:
Cinnamyl Alcohol Dehydrogenase
IFG8898IFG8612
MONOPHYLLA JVS
LONGAEVA JVS
CHIAPENSIS JVS
CHIAPENSIS 02
CHIAPENSIS 01
100
FLEXILIS JVS
FLEXILIS 01S5
FLEXILIS 03S1
AYACAHUITE 04S2
AYACAHUITE 03S2
STROBIFORMIS 01S1
84
62
84
MONTICOLA JVS
MONTICOLA 02S3
70
60
100
65
ALBICAULIS 01S1
ALBICAULIS 03S1
ALBICAULIS 04S1
64
LAMBERTIANA
STROBUS 02S1
STROBUS JVS
STROBUS 03S1100
88
57
86
67
LAMBERTIANA 08S2
LAMBERTIANA 02S1
53
100
Phylogenetic Tree
Phylogenetic Tree
Outgroups
Bootstrap Support (%)
MONOPHYLLA JVS
LONGAEVA JVS
CHIAPENSIS JVS
CHIAPENSIS 02
CHIAPENSIS 01
100
FLEXILIS JVS
FLEXILIS 01S5
FLEXILIS 03S1
AYACAHUITE 04S2
AYACAHUITE 03S2
STROBIFORMIS 01S1
84
62
84
MONTICOLA JVS
MONTICOLA 02S3
70
60
100
65
ALBICAULIS 01S1
ALBICAULIS 03S1
ALBICAULIS 04S1
64
LAMBERTIANA JVS
STROBUS 02S1
STROBUS JVS
STROBUS 03S1100
88
57
86
67
LAMBERTIANA 08S2
LAMBERTIANA 02S1
53
100
Phylogenetic TreeMONOPHYLLA JVS
LONGAEVA JVS
CHIAPENSIS JVS
CHIAPENSIS 02
CHIAPENSIS 01
100
FLEXILIS JVS
FLEXILIS 01S5
FLEXILIS 03S1
AYACAHUITE 04S2
AYACAHUITE 03S2
STROBIFORMIS 01S1
84
62
84
MONTICOLA JVS
MONTICOLA 02S3
70
60
100
65
ALBICAULIS 01S1
ALBICAULIS 03S1
ALBICAULIS 04S1
64
LAMBERTIANA JVS
STROBUS 02S1
STROBUS JVS
STROBUS 03S1100
88
57
86
67
LAMBERTIANA 08S2
LAMBERTIANA 02S1
53
100
Phylogenetic TreeMONOPHYLLA JVS
LONGAEVA JVS
CHIAPENSIS JVS
CHIAPENSIS 02
CHIAPENSIS 01
100
FLEXILIS JVS
FLEXILIS 01S5
FLEXILIS 03S1
AYACAHUITE 04S2
AYACAHUITE 03S2
STROBIFORMIS 01S1
84
62
84
MONTICOLA JVS
MONTICOLA 02S3
70
60
100
65
ALBICAULIS 01S1
ALBICAULIS 03S1
ALBICAULIS 04S1
64
LAMBERTIANA JVS
STROBUS 02S1
STROBUS JVS
STROBUS 03S1100
88
57
86
67
LAMBERTIANA 08S2
LAMBERTIANA 02S1
53
100
Phylogenetic TreeMONOPHYLLA JVS
LONGAEVA JVS
CHIAPENSIS JVS
CHIAPENSIS 02
CHIAPENSIS 01
100
FLEXILIS JVS
FLEXILIS 01S5
FLEXILIS 03S1
AYACAHUITE 04S2
AYACAHUITE 03S2
STROBIFORMIS 01S1
84
62
84
MONTICOLA JVS
MONTICOLA 02S3
70
60
100
65
ALBICAULIS 01S1
ALBICAULIS 03S1
ALBICAULIS 04S1
64
LAMBERTIANA JVS
STROBUS 02S1
STROBUS JVS
STROBUS 03S1100
88
57
86
67
LAMBERTIANA 08S2
LAMBERTIANA 02S1
53
100
Phylogenetic Tree
1
MONOPYLLA JVS
LONGAEVA JVS
CHIAPENSIS JVS
CHIAPENSIS 02
CHIAPENSIS 01
FLEXILIS 01S5
FLEXILIS 03S1
AYACAHUITE 04S2
AYACAHUITE 03S2
STROBIFORMIS 01S1
MONTICOLA JVS
MONTICOLA 02S3
ALBICAULIS 01S1
ALBICAULIS 03S1
ALBICAULIS 04S1
LAMBERTIANA JVS
STROBUS O2S1
STROBUS JVS
STROBUS 03S1
LAMBERTIANA 08S2
LAMBERTIANA 02S1
FLEXILIS JVS
Phylogenetic Tree
1
MONOPYLLA JVS
LONGAEVA JVS
CHIAPENSIS JVS
CHIAPENSIS 02
CHIAPENSIS 01
FLEXILIS 01S5
FLEXILIS 03S1
AYACAHUITE 04S2
AYACAHUITE 03S2
STROBIFORMIS 01S1
MONTICOLA JVS
MONTICOLA 02S3
ALBICAULIS 01S1
ALBICAULIS 03S1
ALBICAULIS 04S1
LAMBERTIANA JVS
STROBUS O2S1
STROBUS JVS
STROBUS 03S1
LAMBERTIANA 08S2
LAMBERTIANA 02S1
FLEXILIS JVS
Future Research:
Continue screening low copy, nuclear genes to compare phylogenetic patterns across the genome. Increase sampling to five individuals from each species that represent the geographic distribution of the species.Extend research to include Asian White Pines.
Special Thanks to:
Howard Hughes Medical InstitutePortland Garden Club
Undergraduate Research, Innovation, Scholarship, and Creativity (URISC)
National Science Foundation
P. strobus