CHROMOSOME 7 THE FRENCH CONTRIBUTION

1. T1112 T1355 T1328 T1428 T1962 T1414 T1497 T1347 T0676 TM18 CT54 TG216 T1257 TG438 T0966 T0731 TM15 T0848 CHROMOSOME 7THE FRENCH CONTRIBUTION SOL 2008 October 12-16, Cologne, Germany

2. INTRODUCTION • Chromosome 7 : • 27 Mbases of gene-dense euchromatin first estimation • 25.1 Mb according to Chang et al, 2008 • Initially 270 BACs to be sequenced (new estimation 250 BACs) • BAC selection: • Selection and validation of the BACs on chromosome • Generation of new tools and resources for rapid selection of the BACs to be sequenced • BAC sequencing: • Private Partner "Cogenics" • From draft production to BAC finishing (Phase 3) SOL 2008 October 12-16, Cologne, Germany

3. BAC selection strategy • Selection and sequencing of 100 "seed BACs" to cover the gene-rich regions of Chromosome 7 • Selection of overlapping BACs by in silico approaches, 3D-DNA pools or macrorray screening. • Check the location of each BAC on Chromosome 7 by FISH and by polymorphism screening on ILs SOL 2008 October 12-16, Cologne, Germany

4. Generation of 3D-DNA pools and Macroarray filters from BAC and Fosmids libraries • These resources were generated in collaboration with the • French Plant Genomic Resource Centre (http://cnrgv.toulouse.inra.fr/) • 3D DNA pools • Half of the HindIII BAC library : 7.8x • The entire MboI BAC library : 7.5x • Available for the community and already widely distributed • (Spain, Italy, UK, USA, , India, Germany) • Macroarray filters • - The entire EcoRI BAC library : 8x • - Half ofFosmid library (150000 including 50000 end sequenced) : 6.1x • Available for the SOL community : the filters can be either sent for in house hybridization or the CNRGV can perform the hybridizations SOL 2008 October 12-16, Cologne, Germany

5. Sequencing Strategy • Until May 2008 : • BAC sequencing using capillary sequencers • Finishing strategy : • Walking on shotgun clones when clone links are available • Direct BAC sequencing between scaffolds • From June 2008: • 454 GS-FLX using Long Paired End Tag reads and Multiplex Identifiers (MIDs) : • First batch of 12 BACs sequenced ( 1/2 run) • 4 BACs directly phase 2 – 1 contig • 7 BACs phase 2 - 2 to 11 contigs • 1 problematic BAC with repeats • results now available on Genbank and SGN (These BAC sequences have been generated with 454 technology) SOL 2008 October 12-16, Cologne, Germany

6. State of the K7 progress - October 2008 16.5 Mbsequenced of which 13.9 Mb are non redundant (15% of redundant sequences) 51 to 55 % of the total euchromatin (27 to 25 Mbases) SOL 2008 October 12-16, Cologne, Germany

7. Contigs of BACs on chromosome 7 • 137 BACs are included in 37 contigs on chromosome 7 1.75 Mb / 22,5 cM 1.13 Mb / 2 cM • 22 BACs remain single SOL 2008 October 12-16, Cologne, Germany

8. Contributors GBF (INRA/INP-ENSAT) Murielle Philippot Pierre Frasse Mohamed Zouine Farid Regad Mondher Bouzayen Cogenics CNRGV Stéphanie Penaud Hélène Bergès Hervé Duborjal Sonia Vautrin Marcel deLeeuw Elisa Prat Diliana Dimova Réjane Beugnot François Pons SOL 2008 October 12-16, Cologne, Germany

9. Details on the Macroarray filters (high density) • 108 plates 384 spotted on 1 membrane (22 x 22 cm) with duplicated spots on a 6x6 pattern. • EcoRI BAC library has been spotted on 2 filters • Fosmids library has been spotted on 4 filters • Image analysis software needed : HDFR3 by INCOGEN SOL 2008 October 12-16, Cologne, Germany