Applying next-generation sequencing to enable marker-assisted breeding for adaptive traits in a homegrown haricot bean (Phaseolus vulgaris L.

Size: px
Start display at page:

Download "Applying next-generation sequencing to enable marker-assisted breeding for adaptive traits in a homegrown haricot bean (Phaseolus vulgaris L."

Transcription

1 Applying next-generation sequencing to enable marker-assisted breeding for adaptive traits in a homegrown haricot bean (Phaseolus vulgaris L.) Andrew Tock Prof Eric Holub & Dr Guy Barker University of Warwick, UK

2 Long-term impact aims Establish molecular breeding capability for adapting Phaseolus bean to the UK climate

3 Long-term impact aims Establish molecular breeding capability for adapting Phaseolus bean to the UK climate Provide UK farmers with a novel, short-season legume break crop that would promote soil renewal and aid black-grass control

4 Long-term impact aims Establish molecular breeding capability for adapting Phaseolus bean to the UK climate Provide UK farmers with a novel, short-season legume break crop that would promote soil renewal and aid black-grass control Establish a food production and supply chain for haricot beans in the UK, providing consumers with a nutritious source of vegetable protein

5 Haricot bean is not currently grown in the UK

6 Crop ideotype Disease resistance (bacterial, viral, fungal)

7 Crop ideotype Disease resistance (bacterial, viral, fungal) Cold tolerance

8 Crop ideotype Disease resistance (bacterial, viral, fungal) Cold tolerance Early maturity

9 Crop ideotype Disease resistance (bacterial, viral, fungal) Cold tolerance Early maturity Root architecture

10 Crop ideotype Disease resistance (bacterial, viral, fungal) Cold tolerance Early maturity Root architecture Nutrient acquisition efficiency

11 Crop ideotype Disease resistance (bacterial, viral, fungal) Cold tolerance Early maturity Root architecture Nutrient acquisition efficiency Plant architecture and growth habit

12 Crop ideotype Disease resistance (bacterial, viral, fungal) Cold tolerance Early maturity Root architecture Nutrient acquisition efficiency Plant architecture and growth habit Human nutritional qualities

13 Crop ideotype Disease resistance (bacterial, viral, fungal) Cold tolerance Early maturity Root architecture Nutrient acquisition efficiency Plant architecture and growth habit Human nutritional qualities Seed colour, size and shape

14 F 6 and F 7 recombinant inbred populations

15 F 6 and F 7 recombinant inbred populations Edmund National Vegetable Research Station cultivar Multiple-disease-resistant Haricot characteristics (Conway et al., 1982)

16 F 6 and F 7 recombinant inbred populations Edmund SOA-BN National Vegetable Research Station cultivar Multiple-disease-resistant Haricot characteristics (Conway et al., 1982) Early maturing Cold-tolerant Drought-tolerant (Dodd and Taylor, 1991, unpublished data)

17

18 Experimental approach Pathology & physiology Identify and characterise contrasting adaptive-trait phenotypes in the parental lines

19 Experimental approach Pathology & physiology Identify and characterise contrasting adaptive-trait phenotypes in the parental lines Derive models of inheritance by characterising and interpreting phenotypic variation within RIL populations: in response to infection with economically important diseases; and with regard to physiological resilience traits

20 Experimental approach Pathology & physiology Identify and characterise contrasting adaptive-trait phenotypes in the parental lines Derive models of inheritance by characterising and interpreting phenotypic variation within RIL populations: in response to infection with economically important diseases; and with regard to physiological resilience traits Computational, molecular & statistical genetics Identify sequence variations between parental lines and develop polymorphic markers using RNA-seq and genotyping-bysequencing (GBS) data

21 Experimental approach Pathology & physiology Identify and characterise contrasting adaptive-trait phenotypes in the parental lines Derive models of inheritance by characterising and interpreting phenotypic variation within RIL populations: in response to infection with economically important diseases; and with regard to physiological resilience traits Computational, molecular & statistical genetics Identify sequence variations between parental lines and develop polymorphic markers using RNA-seq and genotyping-bysequencing (GBS) data Derive a high-resolution genetic map for a bi-parental RIL population

22 Experimental approach Pathology & physiology Identify and characterise contrasting adaptive-trait phenotypes in the parental lines Derive models of inheritance by characterising and interpreting phenotypic variation within RIL populations: in response to infection with economically important diseases; and with regard to physiological resilience traits Computational, molecular & statistical genetics Identify sequence variations between parental lines and develop polymorphic markers using RNA-seq and genotyping-bysequencing (GBS) data Derive a high-resolution genetic map for a bi-parental RIL population Define a mapping interval for potentially durable racenonspecific halo blight resistance

23 Variant-calling pipeline (Bolser, 2014)

24 Candidate-gene SNP and INDEL markers CAPS assay of SNPs Fragment analysis of INDELs

25 Genotyping-by-sequencing (GBS) Reduced genome representation

26 Genotyping-by-sequencing (GBS) Reduced genome representation Genome-wide simultaneous SNP/INDEL discovery and genotyping

27 Genotyping-by-sequencing (GBS) Reduced genome representation Genome-wide simultaneous SNP/INDEL discovery and genotyping Reference-anchored or pairwise alignment of reads (tags)

28 The plant immune system (Dangl et al., 2013)

29 Halo blight Caused by the Gram-negative bacterium Pseudomonas syringae pv. phaseolicola (Psph)

30 Halo blight Caused by the Gram-negative bacterium Pseudomonas syringae pv. phaseolicola (Psph) Type III secretion system

31 Halo blight Caused by the Gram-negative bacterium Pseudomonas syringae pv. phaseolicola (Psph) Type III secretion system Seed-borne

32 Halo blight Caused by the Gram-negative bacterium Pseudomonas syringae pv. phaseolicola (Psph) Type III secretion system Seed-borne Spread by inoculum-splash and wind during rainfall

33 Halo blight Caused by the Gram-negative bacterium Pseudomonas syringae pv. phaseolicola (Psph) Type III secretion system Seed-borne Spread by inoculum-splash and wind during rainfall Bacteria enter through leaf stomata and grow in intercellular spaces

34 Halo blight Caused by the Gram-negative bacterium Pseudomonas syringae pv. phaseolicola (Psph) Type III secretion system Seed-borne Spread by inoculum-splash and wind during rainfall Bacteria enter through leaf stomata and grow in intercellular spaces Cultivated and weedy alternative hosts include most Phaseoleae tribe members

35 Halo blight (CABI, 2012) Causes yield losses of up to 45% (Singh and Shwartz, 2010)

36 Halo blight (CABI, 2012) Causes yield losses of up to 45% Annual losses of thousand tonnes in sub-saharan Africa (Singh and Shwartz, 2010) (Wortmann et al., 1998)

37 Halo blight (CABI, 2012) Causes yield losses of up to 45% Annual losses of thousand tonnes in sub-saharan Africa (Singh and Shwartz, 2010) (Wortmann et al., 1998) Disease prevention Genetic resistance conferred by regionally appropriate R genes

38 Psph race 6 was undetected by known R-genes Table 1. Host reactions of differential cultivars and type accessions when inoculated with isolates of the nine identified races of Pseudomonas syringae pv. phaseolicola, with putative resistance (R) genes indicated Adapted from Teverson (1991: 60) and Taylor et al. (1996a,b: 474, 482), as modified by Miklas et al. (2011: 2440). +, apparent susceptible (compatible) reaction;, apparent resistant (incompatible) reaction; a, apparent resistant reaction with severe hypersensitive response.

39

40

41

42

43

44

45 F 6 S E and F 7 E S RILs; a quantitative trait? Error bars: standard deviation of infection scores recorded following separate inoculations with Psph race 6 isolate 716B (2 4 replicates) and race 1 isolate 725A (2 4 replicates).

46 F 7 and F 13 J. D. Taylor lines; a quantitative trait? Error bars: standard deviation of infection scores recorded following separate inoculations with Psph race 6 isolate 716B (1 replicate) and race 1 isolate 725A (1 replicate).

47

48 Race 6 resistance maps to one major-effect locus P <

49 Race 1 resistance maps to the same locus P <

50 Pse-3 maps to the I gene (BCMV) locus P < P > 0.1

51 LG 1 LG 2 LG 5 LG 6 LG 7 fin (growth habit) Growth habit V (black / violet seed coat, hypocotyl colour) P (potentiates pigment in seed coat, flower & hypocotyl, & pod speckling) Pse-3 (HB) & I (BCMV)

52 Preliminary conclusions and further work Potentially durable race-nonspecific resistance to halo blight is governed by one major-effect locus within a ~600-kbp mapping interval

53 Preliminary conclusions and further work Potentially durable race-nonspecific resistance to halo blight is governed by one major-effect locus within a ~600-kbp mapping interval Key recombinants to be genotyped at parental polymorphisms within noncoding regions to reduce interval size

54 Preliminary conclusions and further work Potentially durable race-nonspecific resistance to halo blight is governed by one major-effect locus within a ~600-kbp mapping interval Key recombinants to be genotyped at parental polymorphisms within noncoding regions to reduce interval size Local Resistance gene enrichment and Sequencing (RenSeq) to be pursued to reveal parental polymorphisms with regard to nucleotidebinding site leucine-rich repeat (NBS LRR) genes located within the mapping interval

55 Preliminary conclusions and further work Potentially durable race-nonspecific resistance to halo blight is governed by one major-effect locus within a ~600-kbp mapping interval Key recombinants to be genotyped at parental polymorphisms within noncoding regions to reduce interval size Local Resistance gene enrichment and Sequencing (RenSeq) to be pursued to reveal parental polymorphisms with regard to nucleotidebinding site leucine-rich repeat (NBS LRR) genes located within the mapping interval Genome-wide association genetics to identify type-iii-secreted virulence effectors conserved amongst all races of the pathovar, as candidate targets for race-nonspecific resistance

56 Acknowledgements Eric Holub Guy Barker Joana Vicente John Taylor Siva Samavedam Peter Walley Laura Baxter Sajjad Awan Vegetable Research Trust Medical and Life Sciences Research Fund