Modern Cowpea Breeding to Overcome Critical Production Constraints in Africa
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1 Feed-the-Future Workshop, PAG XX, Jan. 17, 2012 Modern Cowpea Breeding to Overcome Critical Production Constraints in Africa CRSP and GCP Teams Presented by Jeff Ehlers University of California, Riverside
2 Cowpea -Vigna unguiculata Most widely grown grain legume in Africa >10 M ha (vs <4M ha for common bean) Semi-arid regions (Sudanian-Sahelian belt) Crop of Feed-the-Future Target Agro-ecology Dry grain, fodder, leaves and immature pods eaten African domesticate with millets and sorghum Off to India and other parts of Asia Southern Europe from Roman times still grown in Italy, Greece Brought to the Americas with slave trade- 17 th century (blackeyed pea, cream peas, etc) Main grain legume in Northeastern Brazil Photo courtesy Dr. B.B. Singh
3 Relatives in the Fabaceae Family Choi et al. 2004b
4 Importance Nutritious, high protein food High protein (similar to common bean) Complements cereal-based protein Cropping system health Nitrogen fixation for subsequent crops Heat and drought tolerant Stable production in face of erratic rainfall Adapted to poor soils Especially good at mining Phosphorus Income production and stability First crop to sell (green pods, leaves, fodder) Contributes to diverse rural income streams 4
5 Yield Constraints addressable by breeding Drought Flower thrips Bacterial blight Viruses Striga Root-knot nematodes Macrophomina
6 Why Breeding Resistant Cowpea Varieties Important Most farmers women Low access to inputs sprays, fertilizer Fields with poor soils Pesticides (Insecticides) hazardous Lack of safety precautions Choice of product Mixing, application Container re-use Put the resistance in the seed!
7 CRSP Breeding and training partnerships in semi-arid cowpea zones: ISRA, Senegal/UCR (1980-Present) IRAD Cameroon/Purdue SARI, Ghana/UCR&USDA ( ) INERA, Burkina Faso/UCR (2003-Present) IIA Angola/UCR 2009-Present 200mm 800mm Phenotyping sites -Drought tolerance Emeriti Prof. Anthony Hall pioneered CRSP cowpea breeding work in 1980 in Senegal Long-term conventional breeding partnerships and seed production pipelines developed (Angola-new)
8 Important CRSP varieties developed Cultivar Developer Pedigree Released Key Tolerance & Res Traits Mouride ISRA-Senegal IT81D-1137/ISRA Melakh ISRA-Senegal IT83S /IS CRSP Niebe IRAD-Camroon Vya/IT81D Lori Niebe IRAD-Camroon IT86D-364/IT81D Mid-season Drought Tolerance, weevil, CAbMV Aphid, Bacterial blight, Terminal Drought Cowpea weevil, foliar diseases, CAbMV Cowpea weevil, foliar diseases, CAbMV Apagbaala UCR-SARI Ghana UCR 148-1/IT81D Aphid, Bacterial blight, Striga Marfo-Tuya UCR-SARI Ghana UCR 518-2/Sumbrisogla 2006 Aphid, Bacterial blight, Striga Yacine ISRA-Senegal Melakh/Ndiaga Aw 2005 Aphid, Bacterial blight, CABMV, Drought KVx 421-2J INERA-Burkina KVx /KVx30G Striga, Bacterial blight, drought KVx INERA-Burkina KVx 396//Sakoula/IT85D Striga, bacterial blight
9 CRSP Training & Impacts Trainee Country MSc PhD Post Doc Present Position Joesph Batieno Burkina Faso WACCI PhD student Ousmane Boukar Cameroon Purdue Senior Cowpea Breeder, IITA K.O. Marfo Ghana UC Riverside Senior Cowpea Breeder, deceased Arsenio Ndeve Mozambique UC Riverside PhD student Ndeye-Ndack Diop Senegal UC Riverside Program Leader, GCP Ndiaga Cisse Senegal UC Davis Purdue Senior Cowpea Breeder, ISRA Samba Thiaw Senegal UC Riverside UC Riverside Senior Agronomist, ISRA Abdel Ismail Sudan UC Riverside UC Riverside Principle Rice Scientist, IRRI Wellington Muchero Zimbabwe UC Riverside UC Riverside Scientist, Oak Ridge Ntl Lab
10 Marker-assisted/Modern Breeding Traditional breeding hampered by low trait heritability, sequential evaluation of traits, specialized screening skills/protocols for each trait MAB advantages Near perfect heritability Can be done any time of year Can select simultaneously for several traits Can select individuals for crossing prior to flowering Specialized techniques/environments not needed for every trait during all selection cycles
11 5 Key Tools for Modern Cowpea Breeding Now Available 1. high-throughput genotyping platform Based on Illumina 1536 GoldenGate Assay 2. high quality consensus genetic map 13 RILs, 1,293, 1,107 markers 3. QTL and trait-linked markers Drought tolerance Resistance to flower and foliar thrips, ashystem blight, bacterial blight, root-knot nematodes 4. fingerprints of >600 parental lines 370 IITA/GCP mini-core 230 IITA, West African lines/cultivars 5. outsourced genotyping platform
12 Key Tool No. 1 - High-throughput genotyping platform SNPs from ESTs (gene based) Illumina 1536 SNP fixed array GoldenGate Assay 1375 technically successful SNPs Applied to mapping populations and germplasm Map construction (Key Tool 2); marker-trait associations (Key Tool 3) Applied to germplasm Parental information (Key Tool 4) for breeding, association and diversity analysis
13 Use Platform to Genotype Mapping Populations Population Origin Type Individuals Mapped SNPs CB27 x UCR 779 UCR RIL CB27 x IT97K UCR RIL B x IT84S-2049 UCR RIL Yacine x ISRA RIL CB27 x IT82E-18 UCR RIL Sanzi x Vita 7 IITA RIL CB46 x IT93K UCR RIL TVu14676 x IT84S IITA RIL CB27 x B-1 UCR RIL Dan Ila x TVu-7778 IITA RIL LB30#1 x LB1162 #7 ZAAS-China RIL IT84S-2246 x Mouride UCR F IT84S-2246 x IT93K-503 UCR F
14 Mapping population data summarization Make maps for each RIL - JoinMap map making software MergeMap software to get Consensus Map 1293 individuals from 13 Mapping populations S N P s SNP _0489 TA AA AA AA AA AA AA AA AA 1_0496 AA -- GG AA AA -- AG AG AG 1_0502 GG AA GG AA AA GG GG AG AG 1_0504 TT CC TT CC CC TT TT TT TC 1_0514 CC TT CC TT TC TT CC TT CC 1_0519 CG CC CC GG CC GG CC CG CC 1_0524 CC CC CC CC CC CC CC CC CC 1_0531 CC GG GC GG GC GC CC CC GG 1_0539 TT TC CC TT TT TT CC TC TC 1_0541 TT TT TT TT TT TT TT TT TT 1_0544 TT CC CC TT TT TC TC CC TT 1_0546 AT AA AT TT TT AA AA TT AA 1_0547 AG AA AA AA GG AA AA AG GG 1_0557 GG GG GG GG GG GG GG GG GG 1_0559 TT TT TT TT TT TT TT TT TT 1_0563 TT -- TC TT CC TC TT CC CC 1_0566 TT CC CC CC CC TT CC TC TC 1_0586 AA AT TT TT TT AA TT AT AT 1_0587 CC CC CC CC GG CG CG CC CG 1_0588 AG AG AG AG AG GG AA AA GG 1_0595 CC CC CC CC CC CC CC CC CC 1_0604 AA TT TA AA AA TT TT TA AA 1_0611 AA AC AA AC AA AA AA AA AA
15 Key Tool No. 2 High quality consensus map Muchero et al., 2009, PNAS map very good, based on 6 RILs, 741 individuals New map based on 11 RILs, 2 F4 breeding populations, 1,293 individuals 1107 mapped SNPs, 856 bins 33% more bins 19% more markers improved order Available on Lucas et al The Genome 4:1-11
16 Key Tool No. 3 Markers for traits Biotic stress markers on the map-example: Root-knot nematode resistance 1. RIL mapping population: CB27 (Res) x B-1(Sus) 2. Near isogenic/closely related lines: CB46 Res CB46Null - Sus (BC6 line) Kiawah Res Speckled Purple Hull - Sus 3. Phenotyping Growth Pouch-lab Nematode screening in the field Field Egg mass count Gall score 16
17 Cowpea LG11 QTL Analysis From RIL Phenotype And Genotype LOD Score Traces for Rk QTL Green = Field California summer 08 Red = Growth chamber pouch test Feb. 09
18 Confirmation from Near Isogenic Lines Sus Res Res Sus Resistant lines have same haplotype at nematode locus identified in RIL analysis Confirms Rk gene region near marker 1_0757 Pinkeye Purple Hull-Null Kiawah-Rk CB46-Rk CB46-Null
19 Key Tool No. 3 Markers for biotic res traits
20 Map linked to 600 lines genotyped Map File (1107 mapped SNPs) SNP Linkage Group Distance cm 1_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ Genotype File (600 lines) SNP IT84S IT93K B IT84S _0635 GG GG GG AA 1_1408 GG GG AA GG 1_1383 TT TT -- TT 1_0136 TT TT CC CC 1_0766 AA AA AA AA 1_0465 AA AA AA AA 1_ AA AA 1_0617 GG GG GG GG 1_1437 GG GG GG GG 1_0342 TT TT TT TT 1_0655 AA AA GG AA 1_0963 CC CC CC CC 1_0134 TT TT TT TT 1_0541 TT AA TT TT 1_1146 TT CC TT TT 1_1157 TT CC TT CC 1_0142 AA AA TT AA 1_1339 AA AA AA AA 1_0656 TT TT CC TT 1_0982 GG GG AA GG
21 S N P s Key Tool No. 4 Map-linked Genotypes SNP Linkage Group cm IT84S-2246 IT93K B 1_ GG GG GG 1_ CC CC TT 1_ TT TT AA 1_ CC CC TT 1_ AA AA GG 1_ CC CC TT 1_ TT TT AA 1_ TT TT CC 1_ CC CC GG 1_ GG GG GG 1_ CC CC TT 1_ CC CC TT 1_ CC CC GG 1_ CC CC CC 1_ GG GG GG 1_ TT TT CC 1_ AA AA AA 1_ CC CC CC 1_ CC CC CC 1_ CC CC CC 1_ CC CC CC 1_ GG GG GG >600 lines, varieties, germplasm GGT display of one cowpea LG
22 Key Tool 5: Lower cost, targeted platform for breeding Illumina is fixed array Good for germplasm exploration when polymorphisms unknown Lots of uninformative data points with biparental breeding populations Flexible, single-plex platform for breeding KBiosciences KASPAR SNP Genotyping Platform developed By GCP through collaboration of UCR and KBiosciences Can select any number of markers, e.g. subset of those polymorphic in biparental cross 1052 Illumina platform mapped SNPs successfully converted to KBiosciences KASPAR assay Illumina and KASPAR 99% identical calls DNA extraction and genotyping done by KBiosciences Leaf discs sent in 96-well plates by FedEx, DHL, etc Works fine from Africa, data return via link in weeks
23 Bringing it together with BreedIt Program to choose a marker set for a given cross Can quickly compare cost versus marker density Typically see polymorphic SNPs/cross Many linked-don t need to pay for all Hand selection laborious Input SNP list (from Key Tool #1) Input marker-trait file (Key tool #3) Input parent genotypes (from Key Tool #2, #4) Program Options Choose any 2 parents from 600 genotyped Choose desired cm interval for background selection Choose cm interval for markers flanking traits Outputs a list of SNPs for KASPAR assay
24 BreedIt output file for marker selection SNP LG cm IT93K IT84S-2246 Polymorphic Trait Selected Reason 1_ AA AA 0 0 1_ BB AA 1 1 Every 5 cm 1_ AA AA 0 0 1_ AA AA 0 0 1_ BB AA 1 0 1_ AA BB 1 0 1_ AA BB 1 1 Flanking 1_ BB AA 1 Flower Thrips 1 Trait 1_ BB BB 0 0 1_ AA AA 0 0 1_ AA AA 0 0 1_ BB AA 1 1 Flanking 1_ BB BB 0 0 1_ _ BB AA 1 0 1_ BB AA 1 1 Flanking 1_ BB BB 0 Foliar Thrips 0 1_ BB AA 1 1 Flanking 1_ BB AA 1 0 1_ AA BB 1 0 1_ BB BB 0 0 1_ BB AA 1 0 1_ BB AA 1 0 1_ AA BB 1 0 1_ _ BB AA 1 0 1_ AA BB 1 0 1_ AA 0 0 1_ AA BB 1 0 1_ BB AA 1 1 Every 5 cm
25 Enabling resources for modern cowpea breeding High quality consensus SNP map Genotypes of >600 lines Illumina 1536 and KASPAR highthroughput SNP genotyping platforms BreedIt Software for decision support Trait markers Molecular Breeding Training underway for African breeders
26 Population development Recombina tion Population development Current Focus Marker-assisted backcrossing Working with NARS partners to improve local cultivars Using markers linked to drought, biotic stresses Marker-assisted recurrent selection Developing a MAGIC population 8 way, 300 lines developed to F2 at present 2 nd Recombination cycle 3 rd Recombination cycle IT84S-2246 IT93K F 1 F 2 F 3 Multilocation phenotyping Single seed descent QTL detection estimate marker effects A B C D E F G H F 1 F 1 F 1 F 1 F 1 F 1 F 2 F 3 F 3 F 1 :4 Multilocation phenotyping
27 University of California, Riverside Lucas, Mitchell Muchero, Wellington Pottorff, Marti Wanamaker, Steve Ehlers, Jeffrey D. Roberts, Philip A. Close, Timothy J. Xu, Shizhong Hu, Zhiqiu Thank You National Agricultural Research System (NARS) Cisse, Ndiaga, ISRA-Senegal Drabo, Issa, INERA-Burkina Faso Antonio Chicapa, IIA-Angola Rogerio Chiulele, Eduardo Mondlane U, Mozambique International Institute of Tropical Agriculture (IITA) Nigeria Muranaka, Satoru Boukar, Ousmane Fatokun, Christian 27
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