21/03/2014 Wheat Ini*a*ve Expert Working Group on Durum Wheat Genomics and Breeding EWG-‐DWGB Luigi CaFvelli Roberto Tuberosa Agricultural Research Council, Italy University of Bologna, Italy Wheat Ini*a*ve mee*ng, Obregon, 24 March 2014 Report on the ac*vi*es of the EWG-‐DWGB (c/o Luigi CaFvelli & Roberto Tuberosa, Italy) • Summary of the EWG-‐DWGB agenda • Roadmap for sequencing the durum wheat genome • Update on the durum SNP consensus map • Special issue on durum wheat genomics (Molecular Breeding) • DWGB workshop and Congress “From Seed to Pasta” (EXPO 2015) • Discussion 1 21/03/2014 Call for experts for the EWG-‐DWGB network Present members (79) who have joined the EWG-‐DWGB Argen*na 2 Australia 2 Austria 1 Canada 5 Ethiopia 4 France 3 Germany 2 Hungary 1 Israel 2 Italy 35 Jordan 1 Kazkhstan Mexico Morocco Peru Russia Spain Tunisia Turkey UK USA 1 1 3 1 1 3 1 5 1 4 Objec-ves of the EWG-‐DWGB • Synergize research ac-vi-es within the durum community. • Iden-fy research priori-es in order to – Enhance the selec-on progress – S-mulate interac-ons with the bread wheat community. • Iden-fy the research priori-es for durum wheat genomics. 2 21/03/2014 Relevant topics • Sequencing of the durum wheat genome. • Characteriza*on of tetraploid germplasm resources. • Marker-‐based breeding ac*vi*es. • Genomics of agronomic and quality traits – Stress tolerance – Yield per se – Quality traits – Requirements for the pasta-‐making industries. Wheat I ni*a*ve EWG o n d urum w heat g enomics a nd b reeding Durum wheat: *me to sequence A roadmap toward the sequencing of durum wheat genome 3 21/03/2014 A roadmap in 5 steps 1. 2. 3. 4. 5. WGS on sorted genomes BAC sequencing Assemblies Annota*on Anchoring 1 -‐ WGS on sorted genomes • Fluorescent In Situ Hybridiza-on In Suspension (FISHIS) technology will be used to isolate pure A and B genomes. • Sorted A and B DNA is being used for WGS sequencing to achieve a sequence coverage of >80X in 2x150 paired ends on the HiSeq plaWorm and of >10X in 600 bp, merged paired end reads on the MiSeq plaWorm. • Mate pair library up to 20 Kb and fosmid library of 40 Kb will be generated and used for scaffolding of con-gs. The final objec-ve is a physical coverage of >30X in mate pairs with spans of 5, 10, 20 and 40 Kb. • This approach is currently being implemented with the durum wheat cul-var “Cappelli” in the labs of G Giuliano & S Lucred (ENEA Rome) S Rombauts (VIB-‐Gent) and G Sanchez Perez (Wageningen Univ). 4 21/03/2014 2 -‐ BAC sequencing • To implement the con-g length aher WGS, a BAC-‐pools sequencing approach will be performed with the Langdom BAC library already available. BAC clones will be sequenced in pools of 384 BACs each using Hiseq2500 and MiSeq Illumina plaWorms to achieve an average coverage of 120X for each BAC clone in the pool. • Ideally, about 1,000 384 BAC pools (a haploid genome coverage of approximately 4X) can be recommended. • M Morgante is expected to coordinate this WP. 3 -‐ Assemblies and scaffolding • Three separate assemblies can be performed, one for the BAC pools and two for the WGS data (one for the A and one for the B genome). The BAC will be assigned to the A and B genomes by using the short-‐read data from the WGS sequencing. Con-gs from the BAC pools and the WGS strategy will then be merged. • The scaffolding will be implemented with the long sequences produced by PacBio SMRT technology and by op-cal mapping data (Bionano genomics). • It was agreed that the assembly of the durum wheat genome using op-cal mapping will only be possible when a high-‐quality assembly is produced (i.e. genome coverage >90% and N50 >250 Kb). 5 21/03/2014 4 & 5 -‐ Annota*on and Anchoring • Annota*on, implemented with informa-on from bread wheat and durum wheat RNAseq data (already available, not a limi-ng factor) • Anchoring, Markers and mapping popula-ons – A high-‐quality consensus map will integrate about 30,000 molecular markers including SSR, DArT® and (mostly) gene-‐based SNP markers from the Illumina wheat 90K Infinium arrays. – Genotyping-‐by-‐sequencing or RNAseq-‐GBS will be apply to increase the markers available for anchoring. Funding • ENEA in Italy (c/o G Giuliano & S Lucred), VIB in Belgium (c/o S Rombauts) and Univ. of Wageningen in The Netherland (c/o G Sanchez Perez) have already invested own resources on WGS. • Support to the project has recently been agreed in Italy by CNR (300K Euro, c/o A Stella) and CRA (50K Euro, c/o L Calvelli). • Addi-onal funding support (50K Euro) has been agreed in Turkey (c/o H Budak). • Addi-onal 50K Euro will be contributed by IGA (Udine, c/o M Morgante) . • In-‐kind contribu-ons are also available (mapping popula-ons and markers c/o CRA and Univ. of Bologna). • Other contribu*on are welcome! • A consor*um agreement is in prepara*on. 6 21/03/2014 Funding situa*on 1. 2. 3. 4. 5. WGS on sorted genomes // FUNDED BAC sequencing // PARTIALLY FUNDED Assemblies // PARTIALLY FUNDED Annota*on // Anchoring // MARKER DEVELOPMENT AND MAPPING POPULATIONS Other expected outputs • SRA for durum wheat genomics and breeding. • Facilitate sinergy and enhance complementari-es among groups ac-ve in research/breeding of durum and/or bread wheat. • Catalyze the interest on durum wheat research of as many countries as possible. • A closer collabora-on with the bread wheat community. • Assembly of an SNP-‐based durum consensus map 7 21/03/2014 Special issue in Molecular Breeding Submission of manuscripts on -‐ durum wheat genomics -‐ genomics-‐assisted breeding Tenta*ve agenda 6/30 – Submission of manuscripts 7/31 – Editorial comments returned 8/31 – Revised manuscripts returned 9/15 – Final decision 10/15 – Galley sent to authors 10/31 – Proofed galley returned to Editor Workshop on durum genomics and breeding 8 21/03/2014 Next ac*ons • Mee-ng on 24 March, 18-‐20 pm (Valle Grande) • Nomina-on of the Execu-ve Commilee (12 members) • Following the nomina-on of the Execu-ve Commilee, a Chair and Vice-‐Chair will be elected (via email) • Based upon the results of the ques-onnaire, the Scien-fic Board will drah a Strategic Research Agenda • A third mee-ng will be held in Gatersleben at the end of June just aher the ITMI congress 9
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