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Research Program Genetic Gains (RPGG) Review Meeting 2021: From Discovery to Delivery- Pigeonpea Genomics - Trait discovery & deployment By Rachit K Saxena

  1. Pigeonpea Genomics: Trait discovery & deployment by Rachit K Saxena Senior Scientist- Applied Genomics
  2. Focus areas  Trait discovery:  Development of genomic resources  Molecular markers  Sequencing data  Transcriptome  Genome assembly  Development of genotyping platforms  Mapping of target traits  Identification of markers traits association  High-resolution mapping for candidate genes  Validation of traits associated markers/ candidate genes  Trait deployment:  Markers assisted selection (MAS & EGS: early generation selection)  Markers assisted back-crossing (MABC)  Optimizing novel genomics approaches  Genomic prediction  Haplotypes for candidate genes
  3. Genomic resources Draft genome (Nature Biotechnology 2012) Mt genome (DNA research 2013) HapMap (Plant Biotechnology Journal 2016) WGRS 292 lines (Nature Genetics 2017) Gene expression atlas (J of Exp Botany 2017) SSRs 3,200 SNPs ~16 million GoldenGate 768 SNPs KASPar assays 1,616 SNPs SNP Array 56 K 454 /FLX reads 496,705 TUSs 21,432 Sequencing data ~1,000 linesPangenome (Plant Biotechnology Journal 2020)
  4. 103 pigeonpea lines: 63 released varieties (1960 to 2014) 40 founder lines  Almost similar nucleotide diversity in varieties released in RP1 (θπ = 0.43) RP2 (θπ = 0.48) RP3 (θπ = 0.49) RP4 (θπ = 0.48)  The top six founders contributed 50.8% to the genetic base of the released varieties  Founders T1, T190 and Bahar appeared in 28.6% varieties Limited genetic base of released cultivars  51,201 polymorphic SNPs/Indels Genotyping platforms  High-Density Axiom Cajanus SNP Array (56K SNPs)
  5. Genotyping platforms  Mid-Density Cajanus SNP Array (2 to 3K SNPs)  10 SNPs panel for traits associated markers
  6. High density genetic maps and novel approaches for MTAs  6,818 SNPs mapped in 974cM Genome-wide association analysis
  7. High resolution mapping of restoration of fertility (Rf)  4867 SNP markers on the genetic map derived from ICPA 2039 × ICPL 87119 (F2) population Population size Mapped markers Average inter marker distance LOD PVE % References Genetic map (cM) Genome QTL size (Mb) 188 78-140 SSRs 7.3-3.1 5598311 to 7664779 2 8.9 13.98-24.17 Bohra et al. 2012 186 306 SNPs 3.2 6474381 to 7664779 1.2 8.7 28.5 Saxena et al. 2018 369 4867 SNPs 0.3 7295478 to 7706211 0.41 51.58 45.06 Present study
  8. Traits mapped Hybrid related traits Restoration of fertility Cytoplasmic male sterility Hybrid seed purity Abiotic stress Drought Biotic stress Fusarium wilt Sterility mosaic disease Yield related traits Days to 50% flowering Days to 75% maturity Pods per plant Seed size Plant height Seeds per pod Seed yield per plant Number of primary branches Number of secondary branches Quality trait Seed protein content Other traits Cleistogamy Growth habit Must have traits (3-5 years) Nice to have traits (>5 years) Product Concept Notes (PCN) PCN in pigeonpea PCN1: Medium duration climate resilient varieties, parental lines and hybrids with resistance to diseases and pests PCN2: Mid-early duration climate resilient varieties, parental lines and hybrids with resistance to diseases and pests PCN3: Early duration climate resilient varieties, parental lines and hybrids with resistance to diseases and pests PCN4: Super-early climate resilient varieties, with resistance to diseases and pests PCN1 PCN2 PCN3 PCN4 PCN1 PCN2 PCN3 PCN4 PCN1 PCN2 PCN3 PCN4 PCN1 PCN2 PCN3 PCN4 PCN1 PCN2 PCN3 PCN4 PCN1 PCN2 PCN3 PCN4 PCN1 PCN2 PCN3 PCN1 PCN2 PCN3 PCN1 PCN2 PCN3 PCN1 PCN2 PCN3 PCN4 PCN1 PCN2
  9. Validated markers Mitochondrial gene specific marker in different A4 derived lines Restorer lines and non-restorer lines Growth habit
  10. Diagnostic markers  Successfully converted in to Intertek platform  13 SNPs for Fusarium wilt resistance  12 SNPs for Sterility mosaic disease resistance  One SNP for Days to 50% flowering  Two SNPs for 100 Seed weight  Six SNPs for Restoration of fertility Early generation screening Hybrid purity testing  43 SNPs for 25 hybrids combinations https://excellenceinbreeding.org/module3/kasp SNP based QC panel Dataset used: 8.6million SNPs from elite breeding lines Validation of KASP assays: 40 SNPs from selected 48 SNPs Minimum number of markers for QC: 10 SNPs
  11. Deployment of molecular markers for pigeonpea improvement at NARS and ICRISAT  ~ 104,000 data points (~84K until Q3 2020 + 20K estimated in Q4) have been generated with diagnostic markers  > 8,000 (~6K until Q3 2020 + 2K estimated in Q4) samples tested  Institutes availing facility:  ICRISAT  ICAR- Indian Institute of Pulses Research (IIPR), Kanpur  Agricultural Research Station (ARS) - Tandur, PJTSAU  Institute of Biotechnology, Hyderabad, PJTSAU  Zonal Agricultural Research Station, Kalaburagi, UAS-R  Regional Agricultural Research Station, LAM Farm, Guntur, ANGRAU  Mahatma Phule Krishi Vidyapeeth (MPKV) Rahuri
  12. Modernization of Pigeonpea improvement
  13. Deployment of genomics activities in breeding scheme
  14. Introgression of Fusarium wilt and Sterility mosaic disease resistant QTL in elite pigeonpea cultivars @ ICRISAT and NARS FW SMD Maharashtra BDN711 X Karnataka X ICP 8863, TS3R Madhya Pradesh TJT501, JKM189 X Uttar Pradesh BAHAR X Telangana PRG176 PRG176 Andhra Pradesh LRG41, LRG52 LRG41, LRG52 ICRISAT ICPL88039, UPAS120  ICPL 20096 used as a common source/ donor parent for FW and SMD resistance  ICRISAT has also supplied seeds of ICPL 20096 to partner institutes from a common source
  15. Novel approaches  Family based mapping approach using MAGIC and NAM populations  Early maturity  Protein content  Yield & Yield related traits  New breeding material  Establishment of haplotypes for target traits
  16. Genomic prediction for next generation hybrids and heterotic groups  WGRS of 104 hybrid parental lines  ~70R X 10 A = 700 crosses (F1)  F1s + 104 parents + 14 checks: phenotyped @ two locations  WGRS of 292 lines  2 years phenotyping @ two locations  >10 years historical data 78,210 potential single-cross hybrids  65% higher than the average yield of all hybrids  191% higher than average yield of all inbred lines  Combinations are being tested by ICAR-IIPR
  17. The IMPROVED pigeonpea genome  The contact matrices depict the correction of the draft assembly (left) and the improved assembly (right) using the Hi-C approach Unpublished
  18. Data management  Submitted genomics datasets from the year 2010 to 2020 @ ICRISAT Dataverse  Available at https://cegsb.icrisat.org/openaccess/  Available at http://bms.icrisat.org/ibpworkbench/main  Published sequencing datasets are also available at NCBI
  19. Constraints & possible solutions  Limited collaboration in themes and research programs  Lack of visibility for crop Scientists  Respective Scientist not included in institute’s initiatives (CtEH; USAID-AVISA)  Disproportionate funds for research activities (ICAR-ICRISAT; CRP)  Crop specific Scientists in ONE research program  Less hierarchical organizational structure  Promote interdisciplinary teams across the region  Appropriate funds distribution according to research activities
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