Baga, M.

Bahrani, H.

Chibbar, R.N.

Larsen, J.

English

University of Saskatchewan Research Archive

Genetic analysis of developmental traits associated with enhanced winter survival in autumn-seeded rye (Secale cereale L.)Hirbod Bahrani1, Monica Baga1, Jamie Larsen2, Ravindra N. Chibbar11.Department of Plant Science, University of Saskatchewan, Saskatoon, SK, S7N 5A8, Canada2.Lethbridge Research and Development Centre, Agriculture and Agri-Food Canada, Lethbridge, AB, T1J 4B1, CanadaOutline• Introduction• Phenotyping of traits• Genotyping by sequencing• Association mapping of traits studied • ConclusionsCold acclimation prepares plants for over-winteringhttps://www.usask.ca/agriculture/plantsci/winter_cerealsVernalizationVegetative  Reproductive phase Spring GenotypeWinter genotypehttp://www.fao.org/docrep/006/x8234e/x8234e05.htmGenotype Spring cereals FSI LT50Bonanza barley 115 -7.7Manitou wheat 160 -9.6Gazelle rye 210 -11.6Genotype Winter cereals FSI LT50Dover barley 300 -15.3Novamichurinka durum wheat 370 -18.1Ulianovkia × Kodiak triticale (8X) 480 -22.6CDC Kerstel Wheat 497 -23.3Karkov 22MC wheat 499 -23.5Ulianovkia wheat 530 -24.7Sangaste rye 550 -25.5Kodiak rye 575 -26.5Cougar rye 620 -28.3Frontier rye 735 -33.0Puma rye 735 -33.0Winter Field Survival Index (FSI)https://www.usask.ca/agriculture/plantsci/winter_cerealsDevelopmental traits associated with winter survival• Final leaf number• Prostrate growth• Anthocyanin productionErect Inter mediate ProstrateHypotheses➢ Winter field survival in fall-seeded cereals is an interaction betweenfreezing tolerance and the developmental traits of plants.➢ Genes that affect the developmental traits have a role indetermining the winter survival potential of plants.➢ Determination of winter survival for a panel of 96 rye genotypes.➢ Analyze rye panel for developmental traits associated with winter hardiness. ➢ Genotype rye panel by  Genotyping by Sequencing  (GBS). ➢ Association mapping of rye panel to identify genomic markers associated with winter survival.ObjectivesOrigin of rye genotypes studied 0102030400 20 40 60 80 100No. of Genotype% Survival2016/2017N=9601020304010 20 30 40 50 60 70No. of Genotype% Survival 2014/2015N=5001020304040 60 70 80 90 100No. of Genotype% Survival2015/2016N=50Winter Field Survival (WFS) in different trialsCorrelation coefficients for 50 genotypesWFS % 2016/2017WFS %2015/2016WFS %  2015/20160.65**WFS % 2014/20150.54** 0.44****Significance of Pearson correlation coefficient P ≤0.001Developmental traits• Final leaf number• Prostrate growth habit• Anthocyanin profile• Days to Anthesis• Top internode elongation• Flag leaf area• Plant height• Awn length Greenhouse rye seed productionFinal Leaf Number (FLN) in different trials010203040509 10 11 12No. of Genotype2016/2017N=96Correlation coefficients for 50 genotypesFLN2016/2017FLN2015/20160.80****Significance of Pearson correlation coefficient P ≤0.001010203040508 9 10 11 12No. of Genotype2015/2016N=50020406080Erect Inter mediateProstrateNo. of Genotype2016/2017N=96Prostrate Growth Habit (PGH) in different trials020406080Erect IntermediateProstrateNo. of Genotype205/2016Correlation coefficients for 50 genotypesPGH2016/2017PGH2015/20160.56****Significance of Pearson correlation coefficient P ≤0.001N=50Winter SurvivalFinal Leaf NumberPlant HeightProstrate Growth HabitAwn LengthTop Internode ElongationDays to  AnthesisFinal Leaf Number 0.53**Plant Height 0.15 0.16Prostrate Growth Habit0.41* 0.32 0.20Awn Length 0.17 0.29 0.37* 0.29Top Internode Elongation0.60** 0.59** 0.27 0.28 0.18Days to  Anthesis 0.51** 0.34 -0.27 0.16 0.06 0.31Flag Leaf  Area 0.44* 0.39* 0.006 0.23 0.33 0.48** 0.50**Correlation coefficients between traits 2015-2016 trial**Significance of Pearson correlation coefficient P ≤0.001; *Significance of Pearson correlation coefficient P ≤0.01Winter Survival Final Leaf NumberPlant HeightProstrate Growth HabitAwn LengthTop Internode ElongationDays to  AnthesisFinal Leaf Number 0.84**Plant Height 0.25 0.17Prostrate Growth Habit0.36** 0.28* -0.007Awn Length 0.13 0.21 0.29* 0.22Top Internode Elongation0.68** 0.49** 0.11 0.17 0.33**Days to  Anthesis 0.29* 0.26 -0.22 0.32* -0.07 0.1Flag Leaf  Area 0.64** 0.66** 0.15 0.27* 0.28* 0.61** 0.19Correlation coefficients between traits 2016-2017 trial**Significance of Pearson correlation coefficient P ≤0.001; *Significance of Pearson correlation coefficient P ≤0.010510152025300510152025304 5 12 23 26 27 30 31 32 33 34 36 37 38 39 40 41 42 43Retention time (min)Analysis of anthocyanin in rye genotypesAU0.0000.0050.0100.0150.0200.0250.0300.035Minutes0.00 5.50 11.00 16.50 22.00 27.50 33.00 38.50 44.00 49.50 55.00MusketeerHigh winter survivalLow winter survival• HPLC• C18 column• Mobile phase:A: 5 %  Formic acid B: 22.5 %  Methanol & 22.5 % Acetonitrile• 8 peaks associated with winter hardy genotypes• Cyanidin-3-O-rutinoside• Peonidin-3-Oglucoside26.90 32.8336.5237.8640.78Genotyping by Sequencing  (GBS)• Sequencing libraryconstructionSize selectionAdd adaptors with barcodes• DNA sequencingIon TorrentFastQ files• Isolation of high-quality genomic DNADNA absorbance ratio A260/A280 A230/A260 1.8 - 2 2 - 2.220 ng/µl DNA quantified by PicoGreen23130 bp9416655743612322027564125DNA digestionPstI/MsPIBioinformatics analysesRemove adaptor/barcodes from FastQ files (Trimmomatic)Aligned to rye ref genome (Bowtie)Determine and filter SNPs (SAM, VCF)GenotypeTotal readsMb Average read length bp No. of SNPsRye 357 108 3444Analysis of GBS data by StructureSubpopulation (K)Mean of estimated Ln probable dataWinter Field Survival in sub-populations of rye genotypeHigh LowWinter survival0510152025301 2 3 4 5 6 7No of GenotypeSub-populationSNPs associated with developmental traits by association mapping Developmental traitsSNPs No.2016SNPs No.2017Winter survival 14 16Final leaf number 15 21Prostrate growth habit 7 10Anthocyanin peaks 21 27Days to anthesis 20 13Top internode elongation 24 13Flag leaf area 20 30Awn length 11 14Plant height 20 29WFS (2016,2017) R26QTL cluster associated with developmental traits, winter field survival and anthocyanin productionPH (2017)ANT peak no. 26,38 (2016)TIE (2016)ANT peak no. 26 (2016)FLN (2016)TIE (2016,2017)ANT peak no. 37 (2016)WFS (2017) ANT peak no. 32,36,41 (2016,2017)AWL (2016,2017)ANT peak no. 37,42 (2017)AWL (2016, 2017)FLN (2016)TIE,FLA (2016,2017)FLA (2017)PH (2016)WFS (2016)PH (2017)123457891011Wheat150 KbConclusions➢ Rye genotypes in panel studied show large variation for winter survival➢ Strong correlations between final leaf number, prostrate growth and winter survival➢ Rye genotypes segregated into seven subpopulations from extremely high to very low winter survival➢ Specific Anthocyanin profile associated with winter hardy rye genotypes➢ The region on rye genome was associated with QTL for several developmental traits, anthocyanin production and winter survival AcknowledgmentSupervisors:Prof. Ravindra N. ChibbarDr. Monica BagaCommittee members:Dr. Jamie LarsenDr. Karen TaninoDr. Christopher EskiwDr. Tom Warkentin

Genetic analysis of developmental traits associated with enhanced winter survival in autumn-seeded rye (Secale cereale L.)

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Genetic analysis of developmental traits associated with enhanced winter survival in autumn-seeded rye (Secale cereale L.)

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