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The genome of Fraxinus excelsior
(European Ash)
Elizabeth Sollars, Laura Kelly, Bernardo Clavijo,
David Swarbreck, Jasmin ...
It’s a tough time to be an ash tree
Emerald Ash Borer (Agrilus planipennis)
Ash Dieback (Hymenoscyphus fraxineus)
Workflow
DNA RNA
Genome AssemblyMitochondrial
assembly
Ab initio gene
prediction
Resequencing 37
European trees
Functional...
Workflow
DNA RNA
Genome AssemblyMitochondrial
assembly
Ab initio gene
prediction
Functional
annotation
Gene Sharing
Analys...
We know…
23 chromosome pairs
880 Mbp genome
Selfed
We sequenced…
Wood tissue
200x coverage
Illumina HiSeq + 454
Pairs 200b...
Repeats, Repeats, Repeats
35% of the genome is classed as repeats.
32%
35%
2%
0.01%
12%
18%
1%
LTR Gypsy
LTR Copia
LINE
SI...
Workflow
DNA RNA
Genome AssemblyMitochondrial
assembly
Ab initio gene
prediction
Functional
annotation
Gene Sharing
Analys...
Workflow
DNA RNA
Genome AssemblyMitochondrial
assembly
Ab initio gene
prediction
Functional
annotation
Gene Sharing
Analys...
Gene finding & Annotation
AUGUSTUS with RNA-SeqAUGUSTUS MAKER
Repeat-masked genome + protein alignments
Evidence Modeller
...
Gene Completeness
David Swarbreck, TGAC
ashgenome.org
The open home of the British Ash Tree Genome project
• Genome Assemblies
• Transcriptome assemblies
• JBrows...
Workflow
DNA RNA
Genome AssemblyMitochondrial
assembly
Ab initio gene
prediction
Functional
annotation
Gene Sharing
Analys...
Workflow
DNA RNA
Genome AssemblyMitochondrial
assembly
Ab initio gene
prediction
Functional
annotation
Gene Sharing
Analys...
Gene Sharing
Clustering of orthogroups
using OrthoMCL
• FEXC: Fraxinus excelsior
• MGUT: Mimulus guttatus
• UGIB: Utricula...
Gene Sharing
Clustering of orthogroups
using OrthoMCL
• FEXC: Fraxinus excelsior
• MGUT: Mimulus guttatus
• UGIB: Utricula...
Gene Sharing
Clustering of orthogroups
using OrthoMCL:
• FEXC: Fraxinus excelsior
• CROB: Coffea robusta
• PTRI: Populus t...
Workflow
DNA RNA
Genome AssemblyMitochondrial
assembly
Ab initio gene
prediction
Functional
annotation
Gene Sharing
Analys...
Workflow
DNA RNA
Genome AssemblyMitochondrial
assembly
Ab initio gene
prediction
Functional
annotation
Gene Sharing
Analys...
Resequencing: Diversity Panel
Diversity Panel
In 355 Mb of the reference sequence:
• Reads from 37 trees, ~10x coverage each.
• Total: 28.7 million poly...
Diversity Panel
• SNP tree made using
SNPhylo
• 18.7 million
polymorphic positions
• Filtered set of 14,000
Diversity Panel
• SNP tree made using
SNPhylo
• 18.7 million
polymorphic positions
• Filtered set of 14,000
Diversity Panel
Future Work
• Gene family expansions
– In ash alone
– In woody species
• Diversity panel
– Network analysis
– SNP allele f...
We are the borers…
resistance is futile.
F. excelsior
F. chinensis
EAB resistance
F. mandschurica
EAB and dieback resistan...
Acknowledgments
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The genome of Fraxinus excelsior (European ash)

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Slides presented at the Plant and Animal Genome XXIII conference in San Diego, 11th Jan 2015.

The talk includes assembly and annotation of the ash tree genome, as well as analysis of gene sharing with other plants and an preliminary look at the diversity of European ash trees.

Published in: Science
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The genome of Fraxinus excelsior (European ash)

  1. 1. The genome of Fraxinus excelsior (European Ash) Elizabeth Sollars, Laura Kelly, Bernardo Clavijo, David Swarbreck, Jasmin Zohren, David Boshier, Jo Clark, Anika Joecker, Sarah Ayling, Mario Caccamo, Richard Buggs. @LizzySollars
  2. 2. It’s a tough time to be an ash tree Emerald Ash Borer (Agrilus planipennis) Ash Dieback (Hymenoscyphus fraxineus)
  3. 3. Workflow DNA RNA Genome AssemblyMitochondrial assembly Ab initio gene prediction Resequencing 37 European trees Functional annotation Gene Sharing Analysis
  4. 4. Workflow DNA RNA Genome AssemblyMitochondrial assembly Ab initio gene prediction Functional annotation Gene Sharing Analysis Resequencing 37 European trees
  5. 5. We know… 23 chromosome pairs 880 Mbp genome Selfed We sequenced… Wood tissue 200x coverage Illumina HiSeq + 454 Pairs 200bp – 40 kbp We used… gsAssembler for 454 reads CLC Genomics Workbench SSPACE scaffolder SOAP’s GapCloser We assembled… 875 Mbp in 89,285 scaffolds N50 of 99 kbp 19% in gaps 446 kbp mt. genome 33 complete, 20 incomplete mt. genes
  6. 6. Repeats, Repeats, Repeats 35% of the genome is classed as repeats. 32% 35% 2% 0.01% 12% 18% 1% LTR Gypsy LTR Copia LINE SINE Class II (DNA transposons) Unclassified Tandem repeats Class I: retrotransposons Laura Kelly, QMUL
  7. 7. Workflow DNA RNA Genome AssemblyMitochondrial assembly Ab initio gene prediction Functional annotation Gene Sharing Analysis Resequencing 37 European trees
  8. 8. Workflow DNA RNA Genome AssemblyMitochondrial assembly Ab initio gene prediction Functional annotation Gene Sharing Analysis Resequencing 37 European trees
  9. 9. Gene finding & Annotation AUGUSTUS with RNA-SeqAUGUSTUS MAKER Repeat-masked genome + protein alignments Evidence Modeller 43,298 genes, 59,154 transcripts. Annotated with BLAST and GO terms David Swarbreck, TGAC
  10. 10. Gene Completeness David Swarbreck, TGAC
  11. 11. ashgenome.org The open home of the British Ash Tree Genome project • Genome Assemblies • Transcriptome assemblies • JBrowse • BLAST tool
  12. 12. Workflow DNA RNA Genome AssemblyMitochondrial assembly Ab initio gene prediction Functional annotation Gene Sharing Analysis Resequencing 37 European trees
  13. 13. Workflow DNA RNA Genome AssemblyMitochondrial assembly Ab initio gene prediction Functional annotation Gene Sharing Analysis Resequencing 37 European trees
  14. 14. Gene Sharing Clustering of orthogroups using OrthoMCL • FEXC: Fraxinus excelsior • MGUT: Mimulus guttatus • UGIB: Utricularia gibba • SLYC: Solanum lycopersicum • CROB: Coffea robusta Total orthogroups: 18,870 Laura Kelly, QMUL
  15. 15. Gene Sharing Clustering of orthogroups using OrthoMCL • FEXC: Fraxinus excelsior • MGUT: Mimulus guttatus • UGIB: Utricularia gibba • SLYC: Solanum lycopersicum • CROB: Coffea robusta The largest ash-specific orthogroups are: • Disease resistance proteins • F-box proteins • Expansin-like proteins Total orthogroups: 18,870 Laura Kelly, QMUL
  16. 16. Gene Sharing Clustering of orthogroups using OrthoMCL: • FEXC: Fraxinus excelsior • CROB: Coffea robusta • PTRI: Populus trichocarpa • ATRI: Amborella trichopoda • PITA: Pinus taeda In 11-species comparison: • 4,283 groups shared by all • 7,562 shared by all angiosperms A group of laccase proteins were expanded in the woody species: role in lignin degradation. (ash, pine, & poplar) Laura Kelly, QMUL
  17. 17. Workflow DNA RNA Genome AssemblyMitochondrial assembly Ab initio gene prediction Functional annotation Gene Sharing Analysis Resequencing 37 European trees
  18. 18. Workflow DNA RNA Genome AssemblyMitochondrial assembly Ab initio gene prediction Functional annotation Gene Sharing Analysis Resequencing 37 European trees
  19. 19. Resequencing: Diversity Panel
  20. 20. Diversity Panel In 355 Mb of the reference sequence: • Reads from 37 trees, ~10x coverage each. • Total: 28.7 million polymorphic positions (~ 1 every 12 bp). • Low Frequency Variant Caller in CLC Genomics Workbench 0 5 10 15 20 25 30 Gene SNPs Non-gene SNPs 0 1,000,000 2,000,000 3,000,000 4,000,000 5,000,000 6,000,000 5'UTR SNPs 3' UTR SNPs CDS SNPs Other mRNA & intron 19% 81% 77% 13% 6% 4% 407,911 causing non-synonymous amino acid change 15,689 at splice sites
  21. 21. Diversity Panel • SNP tree made using SNPhylo • 18.7 million polymorphic positions • Filtered set of 14,000
  22. 22. Diversity Panel • SNP tree made using SNPhylo • 18.7 million polymorphic positions • Filtered set of 14,000
  23. 23. Diversity Panel
  24. 24. Future Work • Gene family expansions – In ash alone – In woody species • Diversity panel – Network analysis – SNP allele frequencies among individuals – SNP verification - KASPAR • Optical mapping using BioNano
  25. 25. We are the borers… resistance is futile. F. excelsior F. chinensis EAB resistance F. mandschurica EAB and dieback resistance F. quadrangulata Medium EAB resistance F. americana Dieback resistance F. pennsylvanica Medium dieback resistance The Fraxinus genus
  26. 26. Acknowledgments

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