The Use of Genetic Tools to Improve^ Hardwoods Keith Woeste and Nicholas LaBonte

Progeny tests, provenance trials, clone trials, disease screens, phenology studies, germplasm collections, nursery studies SSRs for genetic diversity

Victory, E. R., Glaubitz, J. C., Rhodes, O.E., and Woeste, K. E. 2006. Genetic homogeneity in nigra () at nuclear microsatellites. Amer. J. Bot. 93: 118-126. Ross-Davis, A. Ostry, M.E., and K. Woeste. 2008. Genetic diversity of butternut (Juglans cinerea) and implications for conservation. Can. J. For. Res. 38 (4): 899-907. Parentage and estimation of relatedness FB30

FA71 N P = 0.0329

0.45 County Road

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0.35 FB 0 500 m FA

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0.25 2012 . 2003 Robichaud, R.L., J.C. Glaubitz, O.E. Rhodes, Jr., and K. Woeste 0.2 BW 2006. A robust set of black microsatellites for parentage 0.15 and clonal identification. New Forests 32: 179 – 196. 0.1 LaBonte, N., M. Ostry, A. Ross-Davis, K. Woeste 2015.

Percent variance among among "families" variance Percent 0.05 Estimating heritability of disease resistance and factors that

0 contribute to long-term survival in butternut (Juglans cinerea 0.1 0.15 0.2 0.25 0.3 0.35 L.). Genetics and Genomes 11: 63-74. Minimum rij threshold 1 1 FA28 FA26 FA13 Gene Flow 11 14 5 28 56 19 1 FA37 27 N Robichaud, R., J. Glaubitz, O.E. 72 Rhodes and K. Woeste. 2010. Genetic consequences of harvest in a mature second growth stand of black FA71 Row walnut ( L.). Ann. For. Crops Sci. 67:702-710. 47 38 Vahdati, K.M., S.M. Pourtaklu. R. Karimi, R. Barzehkar, R. Amiri, M Mozaffari, K. 13 Woeste. 2015. Genetic diversity and gene FA20 County road flow of some Persian walnut populations in southeast of Iran revealed by SSR 28 0 100 m house 19 markers. Systematics and Evolution 301:691:699 Selfs Within-stand matings Immigrant or unassigned matings Forensics and clone identity

Coggeshall, M. and K. Woeste. 2009 Microsatellite and phenological descriptors identify eastern black walnut cultivars in Missouri, USA. Acta Horticulturae International Symposium on Molecular Markers in Horticulture 859 pp. 93-98

McKenna, J.R., W.A. Geyer, K. Woeste, and D. Cassens. 2015. Propagating figured in black walnut. Open J. Forestry 5:518-525. Sequence Data Species identity and differentiation including hybrids

Zhao, P., and K. Woeste. 2010. DNA markers identify hybrids between butternut (Juglans cinerea L.), and Japanese walnut ( Carr.). Tree Genetics & Genomes 7:511-533.

Miller, A., K. Woeste, S.L. Anagnostakis, D.F. Jacobs. 2014. Exploration of a rare population of Chinese in North America: Stand dynamics, health, and genetic relationships. AOB . http://aobpla.oxfordjournals.org/cgi/reprint/plu065? ijkey=3Q3XdABVkHSCm1d&keytype=ref Sequence Data Genome assembly and Gene discovery Q. robur

Q. rubra Chloroplast Q. rubra 161, 304 bp

Q. nigra

Worthen-Alexander, L. and K. Woeste (2014) Pyrosequencing of the northern red (Quercus rubra L.) chloroplast genome reveals high quality polymorphisms for population management. Tree Genetics and Genomes. Broders, K., Woeste, K., San Miguel, P., Westerman, R., Boland, G. 2011. Discovering Single Nucleotide Polymorphisms (SNPs) in 454 sequence data from Ophiognomonia clavigignenti-juglandacearum, an Ascomycete with an uncharacterized genome. Molecular Ecology Resources 11:693-702. Sequence data Process discovery: Heartwood genes

Huang, Z., R. Meilan and K. Woeste. 2009. A KNAT3-like homeobox gene from Juglans nigra L., JnKNAT3-like, highly expressed during heartwood formation. Plant Cell Rep. 28 (11): 1717-1724 .

Huang, Z.; Zhao, P.; Medina, J.; Meilan, R.; Woeste, K. 2013. Roles of JnRAP2.6-like from the transition zone of black walnut in hormone signaling. PLoSOne 8(11): e75857. Published online 2013 November 12. doi: 10.1371/journal.pone.0075857 . Sequence data Ecological genomics and endophyte discovery

James J. Jacobs, Keith E. Woeste, Michael E. Ostry, Surveying Fungal Symbiont Diversity in CastaneaUsing and Charles H. Michler Inhibition of Ophiognomonia High-Throughput Sequencing Data from and Twigs clavigignenti-juglandacearum in vitro by fungi associated with Nicholas R. LaBonte and Keith Woeste butternut, Japanese walnut and hybrid butternut Present / Future: Discovery/ Diversity of genes related to adaptation (Paola, Nick)

By combining data from field studies with detailed insight into gene sequence variation and variation in gene expression we can identify genetic regions and even genes or parts of genes that regulate complex adaptive traits.

Ongoing research Sequence diversity across three blight-resistance QTL in Chestnut. Sequence diversity in ten genes related to bud-break in J. regia.

Model manuscript: Verne et al. 2011. Global transcriptome analysis of constitutive resistance to the white pine weevil in spruce. Genome Biology and Evolution 3:851-867. Present / Future: Genome evolution, comparative genomics, and identification of hybrid barriers

Ongoing research Comparison of the genomes of , Juglans nigra, Juglans cinerea and Juglans cathayensis (Zhao et al.) Phylogeny, biogeography and evolution in the genus Juglans. (Bai et al.)

Model: A De La Torre, P K Ingvarsson and S N Aitken. 2015. Genetic architecture and genomic patterns of gene flow between hybridizing species of Picea. Heredity 115, 153–164. Future: Marker development, including SNP detection and validation (~50,000 markers) Genome assembly of important hardwoods Map assembly and scaffold building

Species Genome size Butternut 700 Mbp Chestnut 794 Mbp Arabidopsis 135 Mbp 1115 Mbp Apple 742 Mbp Poplar 500 Mbp Loblolly 20,000 Mbp

Garvin et al. 2010. Application of single nucleotide Neale, D., et. al. 2014. Decoding the polymorphisms to non-model species: a technical review. massive genome of loblolly Molecular Ecology Resources 10, 915–934 Model pine using haploid DNA and novel Peterson et al. 2012. Double Digest RADseq: An Inexpensive Manuscripts assembly strategies. Genome Method for De Novo SNP Discovery and Genotyping in Model Biology. and Non-Model Species. PLoS One. Future: Comparative transcriptomics (where and when what genes are expressed)

Wachowiak et al. 2015. Comparative transcriptomics of a complex of four European pine species. BMC Genomics. Mittapalli et al. 2010. Tissue-Specific Transcriptomics of the Exotic Invasive Insect Pest Emerald Ash Borer (Agrilus planipennis). PLoS One Cohen et al. 2010. Comparative transcriptomics of drought responses Model in Populus: a meta-analysis of genome-wide expression profiling in mature Manuscripts leaves and root apices across two genotypes. BMC Genomics Future: Fine scale trait mapping and functional genomics—a job for every gene and targets for gene modification

Name Description Type Libraries Chinese Chestnut, Canker associated with CCCanker cDNA Tissues this organism Arild Husby et al. Proc. R. Soc. B 2015;282:20150156 Chinese Chestnut, Mahogany, CCMHS cDNA Healthy Stem

Chinese Chestnut, Nanking, CCNHS cDNA Healthy Stem

Chinese Chestnut, Whole Plant, CCWP1 cDNA Tree 1

Chinese Chestnut, Whole Plant, CCWP2 cDNA Tree 2

CMCMBd Vanuxem ramet from CMCMBd BAC Greg Miller Mah series

CMFMEa CMFME A Flower fraction 678 cDNA CMFMEb CMFME B Flower fraction 9 cDNA CMFMEc CMFME C Flower fraction 10 cDNA CMLMEa CMLME A fraction 567 cDNA CMLMEb CMLME B Leaf fraction 8 cDNA CMRMEa CMRME A Root fraction 567 cDNA CMRMEb CMRME B Root fraction 8 cDNA CMSMEa CMSME A Stem fraction 678 cDNA CMSMEb CMSME B Stem fraction 9 cDNA

CM_MBc chinese chestnut leaf, CM_MBc Vanuxem or Mahogany from BAC Greg Miller

CMCMBb Vanuxem ramet from CMCMBb Greg Mi Model Groover, A. 2005. What genes make a tree? Manuscript Trends in Plant Science. Current /Future: Fine scale genetic diversity studies.

Model Hardy et al. 2006. Fine‐scale genetic structure and gene dispersal inferences in 10 Manuscript Neotropical tree species. Molecular Ecology 15: 559- 571. Future: Genome based selection, genetic architecture of complex traits.

Resende, MDV. et al. 2012. Genomic selection for growth and wood quality in Eucalyptus: capturing the missing heritability and accelerating breeding for complex traits in forest . New Phytologist 194: 116-128. Model Grattapaglia, D. 2011. Genomic selection in forest tree Manuscripts Breeding. Tree Genetics and Genomes 7:241-255. Non-molecular tools for the future Remote sensing and mass phenotyping

Geo-referenced database of all plantings National Forest Inventory and Jones et al. 2009. Thermal infrared imaging of crop Analysis Model canopies for the remote diagnosis and quantification of Spatial Data Services Manuscripts plant responses to water stress in the field. http://www.fs.fed.us/ne/fia/spatial/charter.html Functional Plant Biology 36:978-989. Genetics and genomics of chestnut (Castanea) in the blight resistance breeding program Nick LaBonte- PhD. student

• Restoration of chestnut forests to the eastern depends on developing highly blight-resistant planting stock. • How can genomics help us reach this goal? • Can we investigate other questions about chestnut ecology using genomics? Accelerating the evaluation cycle for chestnut • Evaluating seedlings in their first year would save resources • Early inoculation screens (we tested leaf method) perform poorly • With a dense genetic map, screening could be performed on the basis of genotype • Useq RAD-se to map blight resistance in a chestnut disease resistance breeding population Genomics of blight resistance in Chinese chestnut • Currently the blight resistance breeding program uses only a few Chinese chestnut resistance donors • Is this a liability? • Depends on the nature of coevolution between chestnut and chestnut blight: balancing vs. negative selection. Chestnut genomics can tell us. Genomics of blight resistance in Chinese chestnut

• Analyze sequence diversity in regions associated with blight resistance in 20 + Chinese and hybrids • Use Illumina sequencing to obtain whole-genome sequence for each individual • This approach also allows us to identify markers associated with differences in blight resistance Genomics of blight resistance in Chinese chestnut Ecological Genetics and Genomics Does the hybrid genetic background of backcrossed chestnuts affect the behavior of seed dispersers? Genomics will allow us to Do dispersers prefer American to Chinese chestnut? identify regions of the chestnut genome that affect disperser behavior.

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moorhen.me.uk Investigating the fungal endophyte and epiphyte community of chestnut

Chestnut DNA sequenced from leaves and twigs also contains DNA from fungi living in the plant- we can identify them from small DNA fragments. What is the role of these endophytes in regulating tree health?