Systematically integrating DNA information into breeding: The MAB Pipeline, case studies in apple and cherry Amy Iezzoni January 31, 2013 Cornell MSU Susan Brown Amy Iezzoni (PD) Kenong Xu Jim Hancock Dechun Wang Clemson Cholani Weebadde Ksenija Gasic Gregory Reighard Univ. of Arkansas John Clark WSU Texas A&M USDA-ARS Dave Byrne Cameron Peace Nahla Bassil Dorrie Main Univ. of Minnesota Gennaro Fazio Univ. of CA-Davis Kate Evans Chad Finn Karina Gallardo Jim Luby Tom Gradziel Vicki McCracken Chengyan Yue Plant Research Intl, Carlos Crisosto Nnadozie Oraguzie Netherlands Oregon State Univ. Eric van de Weg Univ. of New Hamp. Alexandra Stone Marco Bink Tom Davis Outline of Presentation The MAB Pipeline Apple skin color Cherry flesh color The MAB Pipeline “Jewels in the Genome” - discovering, polishing, applying QTL discovery MAB Pipelining Breeding (looks promising...) (polishing...) (assembling into masterpieces) Socio-Economics Surveys (example for apple) Washington Michigan Market Breeders Producers Producers Intermediaries Fruit flavor 43 41 23 Fruit crispness 15 23 10 Exterior color 26 Fruit firmness 6 7 5 Shelf life at retail 7 7 3 Sweetness/soluble solids 6 7 3 Sugar/acid balance 9 7 External appearance 13 No storage disorders 7 4 Disease resistance 2 5 Storage life 5 Other fruit quality…2 3 Size 3 Juiciness 2 Tartness Shape Phytonutrient Aroma % of respondents020406080100 Reference Germplasm McIntosh Melba LivelRasp Jolana Williams F_Spartan Spartan PRI14-126 Starr OR38T610 F_Williams NJ53 PRI14-226 Minnesota Delicious KidsOrRed PRI14-644 Cox Gala Sonora NJ8 GoldenDel Winesap F1_9433-2-2 PRI612-1 NJ24 RomBeauty NJ67634 F1_9433-2-8 F_Ill_#2 Mflor821 P21R4A30 Jonathan F2_26829-2-2 NY65707-19 Keepsake Ill_#2 F_P21R4A30 M_MN1764 Prima NJ123249 BP Set Frostbite F_Honeycrisp NJ27 NJ110037 NorthSpy Melrose Honeycrisp Sonya JerseyBla PRI668-100 Macoun Julyred F_Gingergold sibLiberty NY752 F_JamesGr PRI54-12 Liberty Jerseymac Wealthy GingerGold MN1691 (n=160) NY913 F_IngMarie JamesGr Florina F_Delfloki NJ37848 Tetofsky NJ130o PRI47-147 YelTransp IngMarie NJ117637 Longfield PRI610-2 NJ31 NJ88 Raritan DuchOlden NJ12 Monroe Rubin F_PattenGr Mantet Edgewood Elstar PRI1661-1 Vanda PRI855-102 WorcPearm Oriole Nicola RallsJan Sharon NY88 WSU2 Mavis Fireside PRI1661-2 PattenGr StateFair Chinook NY241 BenDavis PRI1018-101 Gravenstein F_Goodland PacQueen Goodland Close M_MolliesDel PacRose PRI643-11 NJ14 AurGoldGa NJ440249 Zestar Jongrimes NJ19237 BC-8S-27-2 Crandall Coop-17 Redfree Wagener M2439 BC-8S-31-56 FrancRus NJ35437 BC-SPA493 PCFW2-134 LadyWill CrownPR Lambourne P7R25A27 NJ136055 BlushiGol Clivia F_Undine Splendour NJ15 Suncrisp Indo Braeburn PRI669-205 PRI2482-100 Grifer Malinda NJ303955 Ambrosia CROP Glockenapfel Cameo Zari MolliesDel DrOldenbu Fuji Bellida PRI1235-100 F_Helios Enterprise Any breeder’s NJ35934 XIIA277 F_EdGoulGol Civni BREEDING F_NJ15 F_Sandow Washington KAZ95-17-14 Ecolette Coop-15 Lundbytrop Cortland NJ109 F_MolliesDel Milwa A28/39 KAZ95-18-07 A849-5 PRI1469-100 Kanzi BeautBath Delcorf KAZ96-05-04 Nicoter Rubinola VKrakers Idared NZ_RGxBB REFERENCE Goldrush additionalAriane Galmac BelHerm Pilot Maigold Priscilla PEDIGREE PRI2059-101 BP SetLaFlam Akane Pinova Rubinstep F_Brina Nevson Sundowner Scarlet CrippsPink Senshu Sansa Rucla Coop-10 NJ381049 Delblush StarkEarl Scifresh Undine PitmPine Linda (n=160)Scilate Delfloki germplasm X4598 Orin Topaz Shizuka MN1764 WSU24 SETS X3174 Toko SET AnanasRei MN1702 WSU37 F_MN1764 Camuzat AutGold Haralson GMAL4591 WSU17 BaumanRei Monark BernRos Telamon GMAL4593 M_Sunrise WSU36 Boskoop Honeygold Delorina GMAL4595 with data BrixPlatt PCF-3-120 Sunrise Calamari Hatsuaki CIVG198 Jonafree WSU38 Calvilla Jonagold Pop_BBxPR Arlet Dalitron CrimsSnow M_NY632 Croncels Gloster Ped-PinxCam NY632 DallaRosa n=480 PRI69-52 ShinGold 4-27-0019 EarlyGold Pop_GaxC15 Alkmene Fiesta Edelb WSU34 Ped-SonxNY241 Helios 5-87-0019 Fiamma 52-05-26 Apollo 25-120-9928 Freedom 5-90-0019 Empire GelEdel Zonga Pop_HCxCP Gewurz Akagi Ped-HCxPit(692) 6-73-0019 Hampshire EdGoulGol Piros Santana HarbRei PRI1956-6 Ped_HCx1764 MN1914 Limoncini Fortune Sandow MPop_HCx1702 4A-100-0013 Magre SchonNord Shinsekai MN1628 MPop_HCxMon Ped-HCxZest GranSmith 4A-38-0013 Napoleone Silken Delgollune GoldenOr Ped-GoldrxNY913 Ontario PermainDor Dolgo Summerfree Merlijn Pop_AurxHC Pristine RedFiled Angold NY75414-1 New York Sabina RenCanada Ariwa Tunda RenChamp CrimsCrisp Discovery WSU46 Robusta5 Belgica WSU35 WSU22 RosBian B_TxBB Rafzubin 4-159-9926 RosdiCal GoldPink RosDop Nicogreen Pop_ArlxCP Brina Ped_HCxJonf RotStett WSU30 BP Set N_PxF Saturn SanLugano Pop_GDxSc Delcoros Sila Creston E_FxGD Stayman Delearly Snowsweet NJ90 E_FxGlo TavBian Ligol TirSpitz Kerr Murray Dalinette (n=160) WeisWint SNP Genome Scans Illumina Infinium® II SNP arrays: developed 2010, used 2011 9K 8K 6K +1K Verde et al. 2012 Chagne et al. 2012 Peace et al. 2012 PLoS One PLoS One PLoS One Standardized Phenotyping • Reference Germplasm Sets Public: 480 individuals Additional: +480 individuals • Standardized phenotyping at multiple locations, esp. for fruit quality (www.rosbreed.org) • Evaluations 2010 - 2012 • Available at Genome Database for Rosaceae (www.rosaceae.org/breeders_toolbox) Pedigree-Based Analysis Pedimap FlexQTLTM for visualizing pedigrees for QTLdiscovery, (with alleles, traits, validation, characterization Identity-By-Descent) RallsJan Delicious W inesap RomBeauty Jonathan M_PRI668-100 GoldenDel F2_26829-2-2 Fuji Crandall PRI14-126 PRI668-100 X-2771 X-3143 X-3318 Flow of a specific linkage block Fruit firmness in apple (HiDRAS data) Jewel Discovery SNP genome scans Pedigree‐ Based QTL discovery Analysis Standardized phenotypic data Systematic QTL Validation – The MAB Pipeline Socio-economics information Breeding markers DNA information PBA: crop-wide PBA: breeding Efficient, program precise, Supported by creative RosBREED’s in-development breeding Breeding Information Management System MAB Successes So Far Two jewels polished in RosBREED, put to routine use in demo breeding programs marker‐assisted parent selection for favorable alleles marker‐assisted seedling selection New generations enriched Routine MAB with QTLs Improved efficiency in seedling evaluation since fall 2010 in RosBREED breeding programs Thousands of inferior seedlings culled - est. $180K saved! Predicted to be (WSU’s apple and sweet cherry breeding programs) large-fruited (& self-fertile) Routine Use of QTLs in Breeding allele 1 allele 2 carriers carriers frequency trait level Use of QTLs: Selecting for favorable QTL alleles enriches for favorable alleles and therefore results in a shift in progeny mean. RosBREED: Filling the Treasure Chest for Rosaceae Breeders Skin Color in Apple: Rf Rf • Alleles = MYB1 promoter 0.00 Mb variations, simple PCR • Alleles 2,3 = non-blushed • Alleles 1, 4 = blushed • Allele 5 = red-fleshed 29.48 Mb Rf = Md-MYB1/MYB10 33.72 Mb can be deduced from published information Rf • Alleles = SNP haplotypes 0.00 Mb from 8K genome scan • Allele “rf” = non-blushed • Allele “Rf” = blushed, dominant 29.48 Mb Rf = Md-MYB1/MYB10 33.72 Mb no. individuals Rf - Predictions “ PercOvrClr” = “% red / overcolor color of skin” Rf rfrf Rfrf RfRf 1 2 3 4 5 no <25% 25-50% 50-75% >75% Rf – Functional Genotypes “ PercOvrClr” = “% red / overcolor color of skin” Rf rfrf Golden Delicious, Granny Smith, Aurora Golden Gala Honeycrisp, Gala, Zestar, Pacific Rose, Braeburn, Pinova, Rfrf McIntosh, Cripps Pink, Splendour, Cameo, Red Delicious RfRfJonathan, Jonamac, Empire, Scired Apple: Skin Coloration Allele & Genotype frequencies in 192 cvs Cover amount over rf rfrf apple skin surface Rf RfRf Rfrf probability 1000 seedlings – how many with score >4? Rfrf x RfRf ---> 647 RfRf x RfRf ---> 818 Marker(s)? 1-3 SNPs from 8K array genome scan Application? Parents: crossing Selections: distinguish Rfrf / RfRf Peach: Maturity Date One trait locus, two ways of genotyping Three 2-SNP haplotypes Two SSR alleles (AA,GA,AG) in four combinations (168,156) in two combinations GA:AA AA:AA AA:AG GA:AG 168:168 156:168 -40 -20 20 +20 +40 -40 -20 20 +20 +40 July days July days Marker(s)? 2 SNPs from 9K array ...or ... 1 SSR Application? Parents: crossing Seedlings: culling Selections: advance What does the raw SNP data look like? (haplotypes = linked alleles that are used to distinguish the different chromosome segments of interest) Ma locus: Most Probable Haplotype Alleles by IBD Courtesy of Sujeet Verma Sujeet Verma @ DC Workshop, 20 Flesh Color in Tart Cherry Flesh color in tetraploid tart cherry Travis Stegmeir & Umesh Rosyara • BACKGROUND – the Myb10 locus controls flesh color in sweet cherry. • Objectives – Determine if the Myb10 locus controls flesh color in tetraploid tart cherry. If so, can DNA information be used to predict color? Flesh Color (visual rating) • Using color cards, an 1 overall flesh color is given. 2 3 4 5 WSU flesh color index card bcde f ghj kl nopu RB_S_3_09729116 BBBBBBBBBBABBB 14 Haplotypes RB_T_3_09782875 A A B A A B B A A B A B A A RB_S_3_10022424 B B B B B A A B B B B B A B RB_S_3_10105783 A B B A A B B B A B A B B B RB_S_3_10162979 BAAAAAAAAAAAAA identified in the RB_S_3_10264563 A A B A A B B A A B A B B A RB_S_3_10573974 ABBAAABBABABBB RB_T_3_10590166 A B B A A B B B A B A B B B RB_S_3_10626205 A B B A A B B B A B B B B B RB_S_3_10675150 A B B A A B B B A B A B A B RosBREED tart RB_S_3_10822211 A B B A A B B B A B A B A B RB_T_3_10908880 A B B A A B B B A B A B B B RB_T_3_12115409 A B B A A B B B A B A A B B RB_S_3_12383977 B A A B B A A A B A B A