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Development of genomics pipelines and its integration with breeding


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  • 1. Development of genomics pipelines and  its integration with breeding hb d Joe Tohme Joe Tohme Workshop in celebration of the lives of  Dr María Jesús ("Chusa") Ginés and Ms Verónica Mera May 12 13, 2010  May 12‐13 2010 Eco-Efficient Agriculture for the Poor
  • 2. Development of genomics pipelines  and its integration with breeding d hb d • Relevant technical advances Relevant technical advances • What are the needs? What are the needs? • What do we have in cassava? What do we have in cassava? • What are the next steps? What are the next steps? Eco-Efficient Agriculture for the Poor
  • 3. A Decade of Major Plant Sciences Breakthrough Whole Genomes Sequencing 2000, July 2000 2002 p g y Citrus pathogen  Xylella  ode p a t ab dops s Model plant Arabidopsis  ce ge o e seque ced Rice  genome sequenced  sequenced in Brazil genome sequenced private public effort Eco-Efficient Agriculture for the Poor
  • 4. Revolution in sequencing technologies Revolution in sequencing technologies 2009‐2010 Routine  Sequenci ng of  plant  genomes  and  associate d pest  Eco-Efficient Agriculture for the Poor
  • 5. Most important plant genomes sequenced   p p g q or about to be sequenced Maize genome sequenced End 2009‐ Cassava sequenced  d d with funds from DOE on  a CIAT genotype a CIAT genotype Jan 2010, three cassava genotypes  sequenced in China in  seq enced in China in collaboration with Brazil Jan 2010, Soybean genome sequenced Eco-Efficient Agriculture for the Poor
  • 6. Sloped land Acid soils, Colombia (Laos) Nigeria Fertile soils, Vietnam Humid environments Amazon, Brazil Dry environments Brazil Orinoco river, Venezuela Eco-Efficient Agriculture for the Poor HERNAN CEBALLOS
  • 7. Cassava Biodiversity Wild Manihot species are  Wild Manihot ild ih i unique sources of genes that  could be used through  g breeding with markers assisted  selection,  and/or genetic transformation and/or  genetic transformation How are going to  How are going to speed up genes  discovery and alleles  discovery and alleles mining for key traits? M. Bonierbale Eco-Efficient Agriculture for the Poor
  • 8. Development of genomics pipelines and  its integration with breeding its integration with breeding Exploit untapped genetic diversity in genebanks, and  breeding populations breeding populations Needs – Access to genomics facilities for gene discovery and  screening – Ph Phenotypic characterizations platform i h i i l f – Development of targeted breeding populations – Bioinformatics capacities – Knowledge of how to design transgenic products and how  to manage the whole process (biosafety and IP) h h l (bi f d ) Eco-Efficient Agriculture for the Poor
  • 9. Cassava Genomics Tools Cassava Genomics Tools Mapping Tools ‐ Several mapping populations linked to breeding l l l k d b d ‐ ~ 350SSRS, 301 AFLP‐RFLPs, 17 Genes ‐ MAS for CMD and Post Harvest Deterioration BAC Libraries BAC Libraries ‐ 3 libraries with 5X, 10X, and 11X coverage ‐ Deposited at Clemson Genome Center Generation of cassava EST Collection ‐ 5,700 Unigene set available − T id tif To identify genes involved in starch metabolism i l di t h t b li − To study cassava ‐ Xanthomonas axonopodis, the causal agent of CBB − Collaborators: University of de Perpignan and IRD, France, U. Nacional Microarray platform developed for cassava yp p A full length cDNA collection of cassava – 20,000 clones sequenced ‐ To develop a large scale of full length cDNA collections ‐ Collaborators: RIKEN Plant Science and RIKEN Genomic Centers, Japan Co abo ato s: a t Sc e ce a d Ge o c Ce te s, Japa Eco-Efficient Agriculture for the Poor
  • 10. Cassava germplasm  Unexplored Source of Novel Traits and Genes Goal: > 30,000 unigenes identified in 2010 Genotype: MTAI16 (KU50) MTAI16 Treatment (Abiotic stress):  ( bi i ) Drought, Heat, PPD, Al toxicity, etc) EST: 20,000 full length cDNA  EST: 20 000 full length cDNA collection Outcome:  First FL cDNA from  Outcome: First FL cDNA from cassava (BMC Plant Biology 2007)  Eco-Efficient Agriculture for the Poor
  • 11. Genomics Tools:  Sequencing and annotations of  Sequencing and annotations of 20000 full length cDNA cassava clones Achievements • A full‐length cDNA library of cassava transcripts to  understand the regulatory networks involved in  stress response  stress response • Gene annotations to identify gene function related  to cassava unique physiological features to cassava unique physiological features • Assist the annotation of the genome sequencing by  DOE‐Joint Genome Institute Eco-Efficient Agriculture for the Poor
  • 12. Sequencing and annotations of  20000 full length cDNA cassava clones 20000 full length cDNA cassava clones Starch Biosynthesis Eco-Efficient Agriculture for the Poor
  • 13. Cassava germplasm  Unexplored Source of Novel Traits and Genes Goal: > 30,000 unigene identified in 2010 Year: 2009‐2010 Whitefly infestation Genotype: MEcu72 (whitefly resistant) Treatment (Biotic stress): Whitefly, Green mites, Mealybugs,       Hornworm, Bacterial blight and pesticide EST: 20,000 FL cDNA collection and 1 m 454 sequence Expected outcome:  novel genes associated with whitefly  MEcu72 resistance Mealybug treatment Year: 2010 Genotype: M. peruviana (wild species), insect resistance Treatment (Biotic stress)  Mealybug, Whitefly, Bacterial  ( ) y g, y, blight and Root rots EST: 20,000 FL cDNA collection  Expected outcome:  novel genes related to insect  resistance CMC40     M. peruviana Eco-Efficient Agriculture for the Poor
  • 14. Global Cassava Genomics Tool Development Sarah Ayling                                       Motoaki Seki Mahidol University Umemura Yoshimi                            Tetsuya Sakurai                 Jarunya Narangajavana Manabu Ishitani                                Minami Matsui           (CASS MOL BIOTECH C di t ) M b I hit i Mi iM t i (CASS‐MOL‐BIOTECH Coordinator) 2009                                                        2010                                                              2011 December: 30,000 FL cDNA  February:  First  Agilent  September: 30,000 gene  March: Marker  sequenced from MEcu72 oligoarray developed  Agilent oligoarray developed for trait of  developed  interest in Thailand March: 20,000 FL cDNA constructed  from M. peruviana (MPER417‐003) March: 1 million read of 454 sequence for  Ecu72 generated E 72 t d Cassava database constructed Eco-Efficient Agriculture for the Poor
  • 15. Xanthomonas axonopodis pv. manihotis Foliar and systemic pathogen F li d t i th Previous studies (Valerie Verdier  Causal agent of Cassava  Bacterial Blight team IRD – CIAT and  Camilo  Lopez, U. Nacional) Lopez U Nacional) • Molecular diversity of populations  in LA and Africa in LA and Africa • Mapping of R genes in cassava  cultivars • Initiate plant‐pathogen interactions Eco-Efficient Agriculture for the Poor
  • 16. Solexa and 454 Sequencing of  Xanthomonas axonopodis pv. Manihotis X h di M ih i Team leader: Adriana Bernal, U. Los Andes, Colombia Team leader: Adriana Bernal, U. Los Andes, Colombia Solexa and 454 Sequencing – Coverage 69,83% with 454 and  33,41%  with Solexa – Assembly used is pseudomolecule  y p (not the real order of the 33  scaffolds) – Still major gaps Still major gaps – 4086 protein‐coding genes  – Many pathogenicity genes common  with other pathovars Eco-Efficient Agriculture for the Poor
  • 17. Gene expression analysis of Cassava Whitefly interaction using  Cassava‐Whitefly interaction using the Cassava Unigene set microarray Tissues from infested and non-infested cassava non infested leaves, from susceptible (CMC40) and resistant (MEcu-72) genotypes at different time during whitefly ( (Aleurotrachelus socialis) attack ) MEcu-72 Resistant genotype Time course experiment Time-course Infested vs non- infested Non-infested Resistant resistant genotype (replica 1, 2 3) Cassava Unigene set micorarray Time-course experiment Eco-Efficient Agriculture for the Poor
  • 18. Genotype used for Sequencing cassava genome CIAT AM 560‐2: line selfed from MCOL 1505  CIAT AM 560 2 li lf d f MCOL 1505 MCOL 1505 has been extensively used by  CIAT breeding project Several derived  lines released in Colombia and Thailand Several derived lines released in Colombia and Thailand MCOL 1505 Self‐pollination S1 Self‐pollination S2 Self‐pollination Hernan Ceballos S3: AM 560‐2 Eco-Efficient Agriculture for the Poor
  • 19. Cassava Database Development  Genotype  information  ts onent Phenotypic  Ph t i information Genome  compo information EST  information Genetic  Genetic DB c information Next activity Genome browser development www cassavagenome org Contact:  Tetsuya Sakurai (RIKEN PSC)  Sarah Ayling  (CIAT)                  Eco-Efficient Agriculture for the Poor
  • 20. Cassava transcripts mapped with reference to the  castor bean, poplar, grape,  and Arabidopsis genome sequences d A bid i Eco-Efficient Agriculture for the Poor
  • 21. Needs for a High throughput SNP platform SNP markers of carotenoid biosynthesis genes for cassava breeding SNP markers of carotenoid biosynthesis genes for cassava breeding ← increase [lycopene] Cloning at Freiburg of carotenoid biosynthesis genes in cassava revealed  Cloning at Freiburg of carotenoid biosynthesis genes in cassava revealed Illuminsa SNP bead Express allelic variation in one (of the two) Phytoene synthase (PSY)  This variation was only found in yellow cultivars and was absent from the  white cassava. SNP ILLUMINA platform being implemented at CIAT white cassava SNP ILLUMINA platform being implemented at CIAT Eco-Efficient Agriculture for the Poor
  • 22. Cassava Tolerance to Postharvest Physiological  Deterioration (PPD)  D t i ti (PPD) In 2009, CIAT evaluated several potentially  tolerant genotypes, scoring the roots 5, 10,  20, and 40 days after harvest Susceptible checks Clones from two different groups of  genotypes showed no symptoms of PPD for  extended days t d dd Consolidates trials planned to provide G X E  data Populations for mapping being developed Eco-Efficient Agriculture for the Poor
  • 23. CIAT Plant Phenotyping Platform Trait Phenotyping Platform Field Phenotyping Platform Discovery/ QTLs /Gene markers Trait  , Gene, QTLs Lead Identification Promising lines Proof of concept Validation  G X E  Feedback to Genomics Back to  Environmental Multilocation trials IS   Impact Impact  Environmental GI Feed B Characterization BMP Eco‐efficient Products Eco-Efficient Agriculture for the Poor
  • 24. CIAT Plant Phenotyping Platform Automation to identify genotypes under water stress  yg yp (>500 lines in 30 minutes) CIAT Customised software High resolution IR camera Barcode reader Eco-Efficient Agriculture for the Poor
  • 25. CIAT Biotechnology  Empowering breeding Empowering breeding Strategy: Biotechnology strongly integrated with breeding and  characterization of germplasm bank characterization of germplasm bank Goal: Agronomical superior genotypes with desirable genes associated  with abiotic, biotic stress, and nutrition Teams working on gene discovery, phenomics, markers assisted  selection, bioinformatics and transgenics Strong focus on very few traits with  well defined target Strong focus on very few traits with well defined target Eco-Efficient Agriculture for the Poor
  • 26. From cassava genomic‐phenomics tools  to breeding b di Challenges now that the genome has been sequenced:  • Allele and allele combinations characterization of breeding  lines for mining allelic diversity and for complex traits lines for mining allelic diversity and for complex traits • High thorough put SNP screening • Urgent needs for bioinformatics capacities and phenomic  platforms Integration of breeding‐genomics tools to  Integration of breeding genomics tools to improve cassava faster Eco-Efficient Agriculture for the Poor
  • 27. Agricultural Sciences for the 21 Century Biotechnology,  Precision  Precision Agriculture Systems Biology Breeding and Synthetic Genomics‐ Biology Bi l Phenomics Bioinformatics and Computational  Biology l Talents, capital, IP management  and public private sector partnerships will be a must Eco-Efficient Agriculture for the Poor