10. a b c a b c a b c a b c A B C A B C A B C A B C X X Parent 3 Parent 4 Parent 1 Parent 2 A B C a b c F1 A B c a B C a B C A B c A b c a b C a b C A b c A b c a B C ABc a b C a B c A b c A b c A b c a B C a B c Bb Bb Bb BB BB BB bb bb bb HEIGHT BB Bb bb GENOTYPE Quantitative Trait Locus Mapping A B C a b c X F1 B b Knott et al. (1997) TAG 84:810-820
16. 2 r2 2n 1n Association Genetics in Conifers Large and Random Mating Population Neale & Savolainen. 2004. Trends in Plant Science. 9:325-330
17. Three approaches to MAS(classified by mapping precision) Modified from Grattapaglia (2007)
18. DNA microarrays to identify genes implicated in the formation of the wood cell wall, through studies of their specific regulation, abundance, or interactions 2000-2003 Expressed genes identification from differentiating xylem (71,377 ESTs in Genbank) QTL mapping Association mapping
19. Phenotype Wood Properties 2001-2004 Water Deficit Resequence SNP Disease Resistance Association Genotype: Illumina- BeadStation 500G-BeadLab Platform, 150,000 data points per week at UCD Genome Center
22. High throughput SNP genotyping in forest trees Pre-SNP era, average marker data points per study ~ 5,000 data points Assume 2,000 studies in total X 5,000 data points ~ 10M data points Last 5 years, forest tree projects at UCD-GC ~ 33M data points
23. wood specific gravity Wood Quality traits microfibril angle S3 secondary wall S2 S1 primary wall cell wall chemistry early late lignin hemicellulose cellulose
26. Water Use Efficiency Stable carbon isotope discrimination in foliage, in two sites (Cuthbert & Palatka). Strong family structure (partial diallel), including 15-24 offspring from 61 families. Cuthbert Palatka
27. FBRC association population in loblolly pine Partial diallel, 15-24 offspring from 61 families. Association with CID (Carbon Isotope Discrimination, related to Water Use Efficiency, in two sites: Cuthbert and Palatka). Analyses using the Quantitative Transmission Disequilibrium Test (QTDT) González-Martínezet al. 2008. Heredity. 101:19-26
37. PineSAP – Sequence Alignment and SNP Identification DNASam – DNA Sequence Analysis and Manipulation Wegrzyn et al. 2009 Bioinformatics 25:2609-2610 Eckert et al 2010 Molecular Ecology Resources 10:542-545 http://dendrome.ucdavis.edu
43. Disease resistanceAssociation Population (409 clones) SNP markers Illumina Infinium: 3938 SNPs for 3100 genes Eckert et al. 2010. Genetics 185: 969-982. Eckert et al. 2010. Genetics 185: 969-982.
44. We couldn’t afford one of those cool PCR robots, so we just got 2 graduate students and a cardboard box. The Cartoon Lab by Ed Himelblau 1055 384-well plates!
45. ADEPT2: Gene Expression Phenotypes: Main Results: 81 SNPs (FDR Q < 0.10) associated to expression for 33 xylogenesis genes: 31 SNPs were nonsynonymous 18 SNPs were synonymous 20 SNPs were intronic 12 SNPs were in UTRs Effect sizes for SNPs in range 1.5-4.5% (r2 from GLM) Most effects were non-additive and due to rare alleles Pleiotropy inferred for 8 genes ΔΔCT values from 112 xylogenesis related genes Palleet al. (2011) Tree Genetics and Genomes. 7:193-206.
46. ADEPT2: Metabolome Phenotypes: Main Results: 61 associations (FDR Q < 0.10) involving 56 SNPs and 44 metabolites. Effect sizes moderate for single SNPs (r2: 4-12%) 292 metabolites from GC-TOF-MS including free amino acids, free fatty acids, sugars and a number of organic acids Statistical Models: Regression on ancestry corrected genotypes and phenotypes for each SNP Bayesian linear mixed models with multiple SNPs and terms for kinship and population structure Eckert et al. New Phytologist (Submitted)
47. ADEPT2: Drought-Tolerance Phenotypes: Main Results: Broad sense heritability 0.4-0.5 Moderate genetic correlations among phenotypes (0.3-0.4). 14 associations detected (FDR Q < 0.05): 6 SNPs with foliar nitrogen 7 SNPs with d13C 1 SNP with height SNP effects small to moderate (GLM: r2 4-9%) Effects largely additive Associated SNPs were mostly to SNPs with low minor allele frequencies (MAF < 0.10). Carbon isotope ratio (d13C), foliar nitrogen content and 2nd year height measured in common garden. BLUPs incorporated spatially autocorrelated errors across the common garden. Statistical Models: Linear mixed and general linear models with and without population structure and kinship corrections for each SNP and trait Cumbieet al. (2011) Heredity Online.
48. ADEPT2: Disease-Resistance Phenotypes: Main Results: 10 associations with small effects for a diverse set of genes Lesion length post infection with Fusarium circinatum collected after 4, 8, and 12 weeks Statistical Models: Bayesian linear mixed models with multiple SNPs and terms for kinship and population structure Quesada et al.(2010) Genetics 186:677-686
51. 7 are of unknown function, predicted proteins, or have no sequence similarity with genes in the databasePeter et al. (unpublished)
52. ADEPT2: Environmental Associations Environmental Gradients: Main Results: 5 associations (FDR Q < 0.10) with small effects mostly with aridity during spring. Eckert et al. 2010. Genetics 185: 969-982. Seasonal aridity gradients across the range of loblolly pine. Statistical Models: Regression on ancestry corrected genotypes and phenotypes for each SNP Ancestry corrections performed via multiple regression and PCA. Eckert et al. 2010. Genetics 185: 969-982.
60. Tree Improvement Infrastructure Tree Improvement Cooperatives: Long-term collaborations with public, private, & academic partners Distributed ownership & responsibilities Goal: to support regeneration activities and decision tools
78. Tree breeders must be trained in the application of genomic breeding technologies
79.
80. Guiding Principles of the Loblolly Pine Genome Project EMPOWERMENT. Our goal is to develop the technologies, platforms and bioinformatics infrastructures to rapidly and inexpensively sequence large and complex genomes of coniferous forest trees. This will allow the forestry community to begin sequencing the many genomes of economic and ecological importance without a dependence on centralized genome centers. ADAPTIVE. We recognize the sequencing technologies are developing rapidly and that we must have the expertise and flexibility to rapidly adopt new approaches into our overall sequencing strategy. COMPARATIVE. We recognize the power of comparative genomics approaches in assembling and annotating genome sequences and will use this approach throughout the project.
81. The pine genome is characterized by diverse and highly diverged sequences Anna S. Kovach1, Jill L. Wegrzyn2, Genis Parra3, Carson Holt4, George E. Bruening5, Carol Loopstra6, James Hartigan7, Mark Yandell4, Charles H. Langley8, Ian Korf3, David B. Neale2,9 1 Genetics Graduate Group, University of California, Davis, CA 95616, USA. 2 Department of Plant Sciences, University of California, One Shields Avenue, Davis, CA 95616, USA. 3 Genome Center, Division of Biological Sciences, University of California, Davis, CA 95616, USA. 4Eccles Institute of Human Genetics, University of Utah, Salt Lake City, Utah 84112, USA. 5 Department of Plant Pathology, University of California, Davis, CA 95616, USA. 6Dept of Ecological Science and Management, Texas A&M University, College Station, TX 77843, USA. 7Agencourt Bioscience Corporation, Beverly, MA 01915, USA. 8 Section of Evolution and Ecology, University of California at Davis, Davis, CA 95616, USA. 9 Institute of Forest Genetics, USDA Forest Service, Davis, CA 95616, USA. Kovach A.S., Wegrzyn J.L., Parra G., Holt C., Bruening G.E., Loopstra C.A., Hartigan J., Yandell M., Langley C.H., Korf I., Neale D.B. (2010) The Pinustaeda genome is characterized by diverse and highly diverged repetitive sequences. BMC Genomics. 11:1-38.
82. Kovach A.S., Wegrzyn J.L., Parra G., Holt C., Bruening G.E., Loopstra C.A., Hartigan J., Yandell M., Langley C.H., Korf I., Neale D.B. (2010) The Pinustaeda genome is characterized by diverse and highly diverged repetitive sequences. BMC Genomics. 11:1-38.
83.
84. Conifer Comparative Genomics Project ( http://dendrome.ucdavis.edu/ccgp ) loblolly pine/Douglas Fir loblolly pine/slash pine loblolly pine/sugar pine
85. “I like trees because they seem more resigned to the way they have to live than other things do” ~ Willa Cather 1913
86. ADAPTATION IN ALPINE CONIFERS David B Neale – UC Davis Elena Mosca, Erica Di Pierro, Nicola La Porta – FEM Giovanni Vendramin – CNR Firenze Piero Belletti – Torino University
87. Introduction Coniferous forests are potentially quite sensitive to climate change Climate change effects: - shift in species ranges to higher elevations due to increase in T Fagus sylvatica L. Pinus mugo - change in forest stand species richness - effects on the interactions among species within the same habitat Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
88. Picea abies Species studied Larix decidua Abiesalba Pinus mugo Aim -effects of climate on conifer population genetics - evidence of local adaptation along environmental gradient Pinus cembra Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
89. CONIFEROUS FORESTS CLIMATE CHANGE EXTINCTION persistence through MIGRATION persistence through ADAPTATION GENETIC DIVERSITY CONSERVATION Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
90. 1. GENETIC DIVERSITY Definition: measure the degree of polymorphism within a population SNP = Single Nucleotide Polymorphism Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
91. Re-sequencing project: Aim: studying the genetic diversity in forest populations Larix decidua Abiesalba Pinus mugo Pinus cembra Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
92. Results: Estimates of genetic diversity: - Watterson’s θ and θπ Genetic diversity: - count of SNP number P. cembra has low genetic diversity Highly adapted In danger ?! Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
93. 2. ADAPTATION Aim: studying the possible interactions between genetic data and environmental factors Methods: ENVIRONMENT DATA SAMPLING PINE NEEDLES GPS DEVICE MODIS/ECA&D TOOLS GENOTYPING CHIP GPS location, Temperature Precipitation… Single Nucleotide Polymorphism DATA ANALYSIS Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
94. 1.Sampling a. Macroscale level: geographical distribution Environmental factors: Elevation Soil Type Expositions Pure/Mixed stands Picea abies/Abies alba Pinus mugo/Pinus cembra Ecological extremes Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
95. A. alba P. abies P. mugo L. decidua P. cembra b. Local scale: Trentino-Alto Adige Provinces - Altitudinal gradient: 2 aspects: North/South 3 plots: high/medium/low elevation 25 trees per plot - Soil gradient: 2 types: lime /silicate soil 2 sides: West/East Adige 65 trees per site -Ecological extremes 25 trees per site - Pure/Mixed stands Picea abies/Abies alba Pinus mugo/Pinus cembra Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
96. On the field - fresh needles collection for each tree In the lab - make the fresh needles dry - DNA extraction Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
97. 2. SNP Genotyping: definition - is the measurement of genetic variations of SNP between species members. Chip Distribution of reaction AA AB BB Fluorescence Data visualization Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
98. 2. Genotyping chip: design SNPs selection : Genotyping chip design: Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
99. AA AB BB Good 3. Data quality checking and final dataset production Bad Finaldataset Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
100. BZ TN 4a. Population structure analysis: STRUCTURE Pritchard et al. Genetics 2000 Picea abies K=4 DISCRIMINANT ANALYSIS of PRINCIPAL COMPONENTS Jombart et al. BMC Genetics 2010 K=3 Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
101. K=3 Abies alba K=7 K=8 Larix decidua K=3 Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
102. K=4 Pinus cembra K=6 K=4 K=4 Pinus mugo Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
103. 4b. Genetic data and geography: A. alba L. decidua 3 2 1 5 4 PCA between geographic areas P. cembra P. mugo Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
104. 4c. Genetic data and climatic data: Multivariate analysis 3 2 A. alba L. decidua 1 4 PCA on these factors: -Lat & Long -Elevation -Aspect -Slope -seasonal T average -seasonal T max and T min -seasonal cumulate P P. cembra P. mugo Impatto dei cambiamenti climatici sulla biodiversita' in Trentino
105. Bayesian analysis Bayenv Coop et al. Genetics 2010 N of SNPs with BF factor > 3 3 2 1 4 MAF and PC2 score A. alba PC2 seasonal T min, T min coldest month, T mean driest quarter
The plot in the lower left gives the sampled counties and population structure estimates for the NCSU population. Colors designate different genetic clusters. The plot on the right is a generic SNP genotyping plot used to call SNP genotypes.
The graph shows the inferred gene network for the targeted genes from Sree’s TGG paper.
The colored matrix gives all pairwise correlations among 292 metabolites. The histogram shows the distribution of the values colored in part A. The plot in the lower right lists in order (top to bottom) of the % phenotypic variance explained for SNPs identified in the Bayesian linear mixed models in a general linear model with population structure covariates. These are the bars. The line gives the # of SNPs identified in the Bayesian linear mixed models with significant effects.
Photo is of the NCSU common garden. The plot shows the spatial variogram across the garden.
Shown in the plot is the distribution of lesion length BLUPs from Quesada et al. 2010. Effect in the table gives the SNP effect/phenotypic standard deviation as a percent. This then gives the effect size scaled to the variation in the phenotype.