INCORPORATION OF    DOWNY MILDEWRESISTANCE IN TARGETED        MAIZE  BREEDING/PARENTAL        LINES HF Galvez, CB Pascual,...
Corn: Philippines’ most importantcereal crop next to riceWithout fungicide, downy mildewseverely affects corn production (...
C8                                   Metalaxyl fungicide and application                  rga01_136.2                     ...
Project Objectives To fine-map the chromosomal locations of likely genes  (QTL) for DMR    To validate the QTLs in multi...
Marker-assisted Purification of Maize Parent LinesPDM Resistant Lines                                  Morphological (roug...
Fine mapping and validation of DMR-QTLValidation of DMR-QTL• Plant materials (Pi23 x P345)   - BC1F1 - genotype data (AMBI...
Establish genetic structure of RIL mapping population                                        118 (Pi23 x P345) BC1F2 famil...
Multi-location disease evaluation for DMR- Natural downy mildew (DM) infection with pre-infected  spreader (sweet corn) pl...
Multi-location disease evaluation for DMR- Final multi-locations trial- 153 BC1F6 lines, parents and sweet corn• Kabacan, ...
RESULTSComparative QTL - UPLB, Isabela and Bukidnonusing (Pi23xP345)BC1F3  - Chromosome 8 (major QTL) – UPLB & Bukidnon  -...
Multilocation QTL analysis of corn RIL population (Pi23 x P345)BC1F6Chrom. Trial site QTL          Flanking           QTL ...
Testcrossing and line conversion for DM resistantmaize variety- DMR-Pi23 x Pi17 and DMR-Pi23- Pi23 – recurrent parent; Pi1...
Marker-assisted introgression of DMRMarker-assisted line purification-2 cycles of morphological data + SSRs (10 maize chro...
Crossing Block: DMR MaizeHybrids/Converted Lines      20 (Pi23 x P345) BC1F6 PDM resistant lines      Pi23 parent (backcro...
Test-Hybrids: Screenhouse DMR Screening and PYT  Initial cross: 4 test hybrids (Isabela and Bukidnon selections)  1301A an...
Conclusion• Corn microsatellite (SSR and EST-SSR) and resistancegene analog (RGA) markers were successfully used to: puri...
MARKER-ASSISTED BREEDING: GENE         PYRAMIDINGPhilippine Downy Mildew Resistance (PDMR)Bacterial Stalk Rot Resistance (...
A BLESSED DAY!
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S2.3. INCORPORATION OF DOWNY MILDEW RESISTANCE IN TARGETED MAIZE BREEDING/PARENTAL LINES

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Presentacion de 11th Asian Maize Conference which took place in Beijing, China from November 7 – 11, 2011.

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S2.3. INCORPORATION OF DOWNY MILDEW RESISTANCE IN TARGETED MAIZE BREEDING/PARENTAL LINES

  1. 1. INCORPORATION OF DOWNY MILDEWRESISTANCE IN TARGETED MAIZE BREEDING/PARENTAL LINES HF Galvez, CB Pascual, AKB Malijan,AO Canama,, RB Quilloy, PH Manguiat
  2. 2. Corn: Philippines’ most importantcereal crop next to riceWithout fungicide, downy mildewseverely affects corn production (80-100% loss)  Metalaxyl fungicide  Pre-treatment of seed materialsPhilippine downy mildew (PDM) in cornis caused by Peronosclerosporaphilippinensis Weston (Shaw)
  3. 3. C8 Metalaxyl fungicide and application rga01_136.2  Human health/environment hazard umc15013.57.6 srga3 umc048  Additional expense or cost in production2.9 asg521.6 1.6 umc1960 1.6 umc1141 Greatly affects seed viability 1.6 umc1728 1.6 umc11495.9  umc089 umc002 9.9 umc01231.0 phi014  PDM resistance reported16.6 umc12010.0 phi125 Use of resistant varieties remains most35.5 rga17_2 effective and economical; resistance sources 177.1 cM available DNA markers mapped in maize genome Chromosome locations of likely genes (QTL) for PDM resistance mapped in P345 x Pi23- maize population
  4. 4. Project Objectives To fine-map the chromosomal locations of likely genes (QTL) for DMR To validate the QTLs in multi-location disease screenings under natural infection of downy mildew To identify resistance gene orthologs and develop PCR- based DNA markers specific to the DMR genes To establish recombinant inbred line population with differential disease reaction to downy mildew To isolate and characterize the cDNA sequences of the DMR genes
  5. 5. Marker-assisted Purification of Maize Parent LinesPDM Resistant Lines Morphological (rouging) then SSR marker purification SSR markers from each maize P345 Nei9008 chromosome Susceptible/Recipient 2 Cycles: bulk then individual plant Inbred Lines marker-assisted purification A line D line E line FG lines HJ lines
  6. 6. Fine mapping and validation of DMR-QTLValidation of DMR-QTL• Plant materials (Pi23 x P345) - BC1F1 - genotype data (AMBIONET, 2000) -BC1F3 - phenotype data (Isabela and Bukidnon)• Method of QTL detection (QTL Cartographer software) - Single marker analysis - Composite interval mapping (CIM)
  7. 7. Establish genetic structure of RIL mapping population 118 (Pi23 x P345) BC1F2 families Select 3-5 phenotypic variants (plants) per family  Plant height/habit  Leaf morphologyP345 (R) Parent Pi23 (S) Parent  Tassel and silk color Bulk and de-bulk SSR analysis for genotype variants  SSR markers from each maize chromosome(Pi23 x P345) BC1F7 seed production 215 (Pi23 x P345) BC1F3, then ear-to- row up to BC1F7
  8. 8. Multi-location disease evaluation for DMR- Natural downy mildew (DM) infection with pre-infected spreader (sweet corn) plants- % disease incidence (% plants infected/line)-period of evaluation – 14, 21, 28, 35, 42 & 49 DAE • Ilagan, Isabela (DA-Isabela Experimental Station) -152 (Pi23xP345)BC1F3 lines - parents and sweet corn as susceptible check • Musuan, Bukidnon (Central Mindanao University) -197 BC1F3 lines - parents and sweet corn as susceptible check
  9. 9. Multi-location disease evaluation for DMR- Final multi-locations trial- 153 BC1F6 lines, parents and sweet corn• Kabacan, North Cotabato (Univ. of Southern Mindanao) - use of reported virulent DM isolate (Carmen)• Banga, South Cotabato (ACM Genetics) - collaboration with private seed producer
  10. 10. RESULTSComparative QTL - UPLB, Isabela and Bukidnonusing (Pi23xP345)BC1F3 - Chromosome 8 (major QTL) – UPLB & Bukidnon - Chromosome 9 – Isabela only - Chromosome 1 – UPLB and Isabela - Chromosme 4 - UPLB only
  11. 11. Multilocation QTL analysis of corn RIL population (Pi23 x P345)BC1F6Chrom. Trial site QTL Flanking QTL LRb Genetic Effects R2(%)c No. detected marker positiona Additive Dominance 1 Isabela rga01_2 –umc67 0.3054 15.93 -6.14 1.15 13.49 2 Bukidnon csu109-bnl8.44 0.2505 11.71 -9.47 0.53 14.90 South Cotabato csu54-umc055 1.1588 20.75 -0.06 - 23.59 3 North Cotabato umc121-phi36 0.0001 11.64 -5.48 0.03 8.92 5 North Cotabato phi91-phi116 2.3954 31.82 -9.87 0.08 29.86 South Cotabato bng1386 - 1.9283 16.21 0.057 61.25 20.4 umc104_1 6 Los Baños umc65-umc59 1.3699 11.79 10.75 4.55 9.88 7 North Cotabato rga16-rga19 0.9461 12.86 -6.19 0.16 11.64 8 Los Baños rga01_1-umc150 0.0001 20.58 20.95 7.67 52.69 9 Isabela umc105-umc113 1.278 24.28 8.32 1.27 24.77 South Cotabato umc81-phi61 0.5471 22.13 -0.081 78.33 25.2 a Position of QTL based on composite interval mapping (CIM) b Peak value of the maximum-likelihood-ratio(LR) test statistic observed for the QTL c Proportion of phenotypic variance explained by the QTL
  12. 12. Testcrossing and line conversion for DM resistantmaize variety- DMR-Pi23 x Pi17 and DMR-Pi23- Pi23 – recurrent parent; Pi17 – other parent for hybrid• Initial set of candidate DMR-Pi23 lines - molecular, field DMR (Isabela) & nursery DMR (UPLB) - 5 BC1F6 lines
  13. 13. Marker-assisted introgression of DMRMarker-assisted line purification-2 cycles of morphological data + SSRs (10 maize chromosomes)Genuine hybrids (SSR markers) - Initial SSR analysis, 87% confirmed genuine hybrids No. of No. of Total SSR No. of true F1 Entry heterozygous homozygous loci screened hybrid plants SSRs SSRs P345 x Pi17 10 10 0 2/2 P345 x Ca00314 10 9 1 14/19 Nei9008 x Ca00314 10 7 3 31/33
  14. 14. Crossing Block: DMR MaizeHybrids/Converted Lines 20 (Pi23 x P345) BC1F6 PDM resistant lines Pi23 parent (backcross for line conversion) Other maize parental line/s: good SCA for test-Hybrid cross
  15. 15. Test-Hybrids: Screenhouse DMR Screening and PYT Initial cross: 4 test hybrids (Isabela and Bukidnon selections) 1301A and 1304B candidate hybrids:  Screenhouse DMR and preliminary agro-morpho (yield) MAS for QTL markers  Had 0% DM incidence 16 more test hybrids for evaluation
  16. 16. Conclusion• Corn microsatellite (SSR and EST-SSR) and resistancegene analog (RGA) markers were successfully used to: purify parent lines and hybridization crosses to incorporate DMR derived from P345 and Nei9008 Re-establish genetic structure of RIL population for genetic mapping and in combined MAS breeding schemes• Resistance QTL were validated in multi-location DMscreening. The major QTL in Chrom 8 was detected inUPLB. The mapping of multiple and location specificQTL suggests variation among DM isolates• Best performing DM resistant lines (Pi23xP345)BC1F7and hybrids (1301A and 1304B) were identified and willbe released as a registered new corn variety and/orgenetic stock.
  17. 17. MARKER-ASSISTED BREEDING: GENE PYRAMIDINGPhilippine Downy Mildew Resistance (PDMR)Bacterial Stalk Rot Resistance (BSRR)Quality Protein Maize (QPM)
  18. 18. A BLESSED DAY!

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