1. Sunflower Genome Consortium Annual Meeting Nicolas Langlade INRA Toulouse Sunflower Genome Consortium San Diego, January 15 2014

2. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 BAC libraries o Preparation CNRGV (H. Bergès, A. Bellec) o BAC-end sequencing LIPM Ultra High Density Genetic map – SFP using Affymetrix microarrays – Genotyping by sequencing – AXIOM map Genome annotation – Transcriptome sequencing – Transcriptome annotation – Genome annotation per se J. Gouzy, S. Carrère, N. Langlade, S. Muños, P. Vincourt

3. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 BAC libraries Preparation -CNRGV (Centre National de Ressources Génomiques Végétales) H. Bergès, A. Bellec 3 BAC librairies HA412-HO BAC-end sequencing -LIPM J. Gouzy, P. Vincourt, S. Muños, N. Langlade, B. Mayjonade Development of a method to perform high-troughput BAC-end sequencing First experiment: BAC-ends are there but too much contamination

4. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 Ultra High Density Genetic map – Genotyping SFP using Affymetrix microarrays P. Vincourt, S. Muños, Y. Lippi, N. Langlade, B. Mayjonade – 105 RILs from XRQxPSC8 population – >3000 mkrs – ~6000 cM  Not used

5. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 Ultra High Density Genetic map – Genotyping by sequencing P. Vincourt, S. Muños, N. Langlade, B. Mayjonade 2 enzymes: PstI and MseI 2 populations: XRQxPSC8 ~192 RILs FUxPAZ2 ~96 RILs Sequencing finished end December 2013 Results expected spring 2014

6. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 Ultra High Density Genetic map – Genotyping using Affymetrix AXIOM P. Vincourt, S. Muños, N. Langlade, B. Mangin In collaboration with Biogemma and Syngenta 197,914 SNPs mostly in expressed sequences from a small panel of elite lines 2 populations XRQxPSC8 180 RILs FUxPAZ2 87 RILs  35,562 polymorphic between XRQ and PSC8

7. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 Ultra High Density Genetic map – Genotyping using Affymetrix AXIOM 1.35,562 polymorphic between XRQ and PSC8 + 1,691 Infinium markers from previous map (Cadic et al., 2013) assign the LG 2. Carthagene Number of markers: 31,757 on the 17 LG Total genetic distance of 1,487.7 cM Number of loci: 1,861 = map XRQxPSC8

8. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 Ultra High Density Genetic map – Genotyping using Affymetrix AXIOM 1.28,529 polymorphic between XRQ and PSC8 + use of AXIOM XRQxPSC8 map to assign the LG 2. Carthagene Number of markers: 17,901 on the 17 LG Total genetic distance of 1,425.3 cM Number of loci: 807 = map FUxPAZ2

9. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 Ultra High Density Genetic map – Genotyping using Affymetrix AXIOM 1.7,076 markers polymorphic between XRQ and PSC8, and between FU and PAZ2 + use of AXIOM XRQxPSC8 map to assign the LG 2. Carthagene Number of markers: 7,076 on the 17 LG Total genetic distance of 1,471.1 cM Number of loci: 1,113 = map consensus XRQxPSC8 & FUxPAZ2

10. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 Ultra High Density Genetic map – Genotyping using Affymetrix AXIOM 1.All markers polymorphic between XRQ and PSC8, or between FU and PAZ2 + use of consensus XRQxPSC8 & FUxPAZ2 map as a skeleton 2. Carthagene Number of markers: 45,566 on the 17 LG Total genetic distance of 1,794.19 cM Number of loci: 2,711 = final map XRQxPSC8 & FUxPAZ2

11. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 Ultra High Density Genetic map – Genotyping using Affymetrix AXIOM 1.Final map XRQxPSC8 & FUxPAZ2 2.Context sequences of the 31,757 markers polymorphic between XRQ and PSC8 and mapped 71bp context sequences Collaborative project with Biogemma and Syngenta Available now

12. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 BAC libraries o Preparation CNRGV (H. Bergès, A. Bellec) o BAC-end sequencing LIPM Ultra High Density Genetic map – SFP using Affymetrix microarrays – Genotyping by sequencing – AXIOM map Genome annotation – Transcriptome sequencing – Transcriptome annotation – Genome annotation per se

13. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 Transcriptome sequencing – Organ-specific transcriptomes XRQ/B inbred line Webportal www.heliagene.org/HaT13l

14. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 Transcriptome sequencing – Suggestion: Range of abiotic stresses on the XRQ/B line: 1.Drought in leaves 1.6-7 week old plants 2.Leaves 3.Range of stress intensities 2.Salt stress 1.Plantlets (10 days) 2.Leaves and Roots 3.2 conditions (CTL, NaCl 100 mM) 3.Osmotic stress 1.Plantlets (10 days) 2.Leaves and Roots 3.2 conditions (CTL, NaCl 100 mM) 4.? Cold 1.Plantlets (10 days) 2.Leaves and Roots 3.4 conditions (CTL time 1, 4°C time 1, CTL time 2, 20°C recovery time 2)

15. INRA participation Sunflower Genome Consortium San Diego, January 15 2014 BAC libraries o Preparation CNRGV (H. Bergès, A. Bellec) o BAC-end sequencing LIPM Ultra High Density Genetic map – SFP using Affymetrix microarrays – Genotyping by sequencing – AXIOM map Genome annotation – Transcriptome sequencing – Transcriptome annotation – Genome annotation per se

16. Principle of a MAGIC population Bandillo et al. Rice 2013 6:11 doi:10.1186/1939-8433-6-11 F1 IM1 IM2 IM2-S1 IM2-S2 IM2-S3 Sunflower Genome Consortium San Diego, January 15 2014

17. Sunflower MAGIC population proposal Two populations – Male 400 RILs – Female 400 RILs Wild parents introgressed in cultivated germplasm – 2*5 different Helianthus species and wild H. annuus from UBC germplasm – 6 different B-lines from collaborators – 6 different R-lines from collaborators Sunflower Genome Consortium San Diego, January 15 2014

18. Tentative list of the wild material involved 10 wild founders from UBC germplasm W1H.annuusOklahoma FT annuus W2H.annuusModern weedy annuus W3H.annuusUtah annuus (sweep for drought) W4H.annuus? W5H.petiolarisGSD Large seeded W6H.neglectusLarge seeded sand dune W7H.argophyllusShort day W8H.bolanderiNorthern range serpintine W9H.anomalus? W10?? Sunflower Genome Consortium San Diego, January 15 2014

19. MAGIC population development W1 X->F1-1 E1 X->IM1-1 E2 x->F1-2 E3 W2 X->F1-3 E2 X->IM1-2 E3 x->F1-4 E4 W3 X->F1-5 E3 X->IM1-3 E4 x->F1-6 E5 W5 X->F1-9 E5 X IM1-5 E6 X->F1-10 E1 … Sunflower Genome Consortium San Diego, January 15 2014

20. MAGIC population development W1 X->F1-1 E1 X->IM1-1 E2 x->F1-2 E3 W2 X->F1-3 E2 X->IM1-2 E3 x->F1-4 E4 W3 X->F1-5 E3 X->IM1-3 E4 x->F1-6 E5 W5 X->F1-9 E5 X IM1-5 E6 X->F1-10 E1 … IM2-1 IM2-2 IM2-3 Sunflower Genome Consortium San Diego, January 15 2014

21. MAGIC population development W1 X->F1-1 E1 X->IM1-1 E2 x->F1-2 E3 W2 X->F1-3 E2 X->IM1-2 E3 x->F1-4 E4 W3 X->F1-5 E3 X->IM1-3 E4 x->F1-6 E5 W5 X->F1-9 E5 X IM1-5 E6 X->F1-10 E1 … IM2-1 IM2-2 IM2-3 IM1-1 IM1-2IM1-3IM1-4IM1-5 IM1-1 IM2-1 IM2-2 ……… IM1-2 ……… IM1-3 …… IM1-4 … IM1-5 Sunflower Genome Consortium San Diego, January 15 2014

22. MAGIC population development W1 X->F1-1 E1 X->IM1-1 E2 x->F1-2 E3 W2 X->F1-3 E2 X->IM1-2 E3 x->F1-4 E4 W3 X->F1-5 E3 X->IM1-3 E4 x->F1-6 E5 W5 X->F1-9 E5 X IM1-5 E6 X->F1-10 E1 … IM2-1 IM2-2 IM2-3 IM2-1-S1-1 IM2-1-S1-2 … IM2-1-S1-20 IM1-1 IM1-2IM1-3IM1-4IM1-5 IM1-1 IM2-1 IM2-2 ……… IM1-2 ……… IM1-3 …… IM1-4 … IM1-5 Sunflower Genome Consortium San Diego, January 15 2014

23. W1 X->F1-1 E1 X->IM1-1 E2 x->F1-2 E3 W2 X->F1-3 E2 X->IM1-2 E3 x->F1-4 E4 W3 X->F1-5 E3 X->IM1-3 E4 x->F1-6 E5 W5 X->F1-9 E5 X IM1-5 E6 X->F1-10 E1 … IM2-1 IM2-2 IM2-3 IM2-1-S1-1 IM2-1-S1-2 … IM2-1-S1-20 40 families Each 20 lines  400 lines 5204005 wilds 6 R-lines Sunflower Genome Consortium San Diego, January 15 2014