Plant genomics and proteomics: applications and i i i C di
Transcript of Plant genomics and proteomics: applications and i i i C di
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Plant genomics and proteomics: applications and i i i C diopportunities in Canadian crops
Faouzi BekkaouiFaouzi Bekkaoui
Rendez-Vous Protéomiqe 2008M t l 10 A il 2008Montreal, 10 April 2008
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Outline
• Overview of crop genomics projects p g p j
Genome Canada projects
Federal Genomics Initiatives
• Examples of proteomic applications
Brassica napus (canola)Brassica napus (canola)
• Research opportunities for future projects
Position paper and potential papersPosition paper and potential papers
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Value of Canadian crops
• Agriculture and food is the fourth largest industry in Canada
• Canadian agriculture will export 28.8 million tonnes of i d il d i 200 h $13 8 billigrains and oilseeds in 2007, worth $13.8 billion.
Canada is one of the world’s largest exporters of agricultural products including flax, canola, lentils andagricultural products including flax, canola, lentils and durum wheat.
• Wheat crops form a $5 billion portion of agricultural production in Canada.
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Importance of Canola
C l t ib t $11 billi t th• Canola contributes over $11 billion to the Canadian economic activity
• More than 52 000 Canadian farmers grow canola• More than 52,000 Canadian farmers grow canola• Canola exports represent 75% of our annual
production and bring over $2 billion back to our p gCanadian economy
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Genome Canada Projects
• I. Functional Genomics of Abiotic Stress, Bill Crosby, U of Windsor, Genome Prairieof Windsor, Genome Prairie
• II. Enhancing Canola Through Genomics, Wilf Keller, g g , ,NRC‐PBI, Genome Prairie
• II. Functional Genomics of Arabidopsis, John Coleman, University of Toronto, Ontario Genomics Institute
• II. The Canadian Potato Genome Project, Barry Flinn, U i it f N B i k & Sh RUniversity of New Brunswick & Sharon Regan, University of Carleton, Genome Atlantic
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Genome Canada Projects
• III. Designing Oilseeds for Tomorrow’s Markets, Randall Weselake ,U of Alberta and GopalanRandall Weselake ,U of Alberta and Gopalan Selvaraj, NRC‐PBI, Genome Alberta
• III. Use of Genomic Tools for Crop Improvements in Temperate Climates, D. Brian Fowler, University of Saskatchewan, Genome Prairie
• GrapeGen ‐ A Genomic Approach to the Identification of the Genetic and EnvironmentalIdentification of the Genetic and Environmental Components Underlying Berry Quality in Grapevine, Steven T. Lund, University of British Columbia
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Federal Genomics Initiatives
NRC‐Genomics and Health Initiative
• focus on canola (Brassica napus)focus on canola (Brassica napus)
– Seed Development
– Seed Metabolism
– Seed Quality
AAFC Crop genomics initiatives
• focus on canola, wheat, soybean and corn
– Disease and insect resistance
T l t t h ld d d ht– Tolerance to stresses such as cold and drought
– Enhanced quality attributes
Development of genomics resourcesDevelopment of genomics resources
• ESTs, DNA arrays, BAC libraries, mutagenized populations etc.
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Arabidopsis thaliana vs Brassica napus
Chromosomes n=5 n=19
Genome size 130 Mb 1200 MbGenome size 130 Mb 1200 Mb
Expressed genes 27,000 80,000-120,000
DNA Homology 85%
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Brassica Seed
SEED COAT
O
SHOOT APEX
HYPOCOTYL
EMB
RYO
COTYLEDON
ENDOSPERM
RADICLE
C t % ENDOSPERMComponents %
Oils 45
Proteins 22
Carboh drates 20Carbohydrates 20Water 9
others 4
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Brassica Seed
SEED COAT
EMB
RYO
SHOOT APEXHYPOCOTYL
COTYLEDON
ENDOSPERM
RADICLE
ENDOSPERM
Example of targets:p g• Enhanced oil content• Modified proteins• Modified oilsModified oils• Reduced anti-nutritional factors
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Seed storage proteinsSeed sto age p ote s
Inhibition of Seed Storage ProteinsHegedus et al g
Gene expression and organisationSharpe et al
Agri-Food Canada Agroalimentaire CanadaAgriculture and Agriculture et
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B napus seed storageB. napus seed storage proteins
Napin (2S albumin) and Cruciferin (12S globulin)p ( ) ( g )
Major types of seed storage proteins in B. napus
20% and 60% of total protein in mature seed
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Protein Production in SeedsProtein Production in Seeds
Basic Requirements to Realize this Potential:Basic Requirements to Realize this Potential:
C i hi h d f• Create space within the seed for new proteins
• Accumulate protein in a location where: p– they are functional
– protected from degradation
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protected from degradation
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The “Empty” Seed ConceptThe Empty Seed Concept
NapinCruciferin
Reduceor80% Cruciferin
OleosinDehydrin
orEliminate Replace
New ProteinsWith
More Utility or Value
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More Utility or Value
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Creating Space for New ProteinsCreating Space for New Proteins
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No ill‐Effect of Storage Protein Depletion
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Gene expression and
• 24 B napus napin genes 4 families
organization in B. napus• 24 B. napus napin genes - 4 families• 18 B. napus cruciferin genes - 4 families• 2 closely related cruciferin genes are most2 closely related cruciferin genes are most
highly expressed• 2 divergent napins are most highly expressed• B. napus cruciferins have similar physical
organisation to Arabidopsis orthologue• B. napus napins have different physical
organisation to Arabidopsis orthologues
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Brassica Seed Developmentand Metabolismand Metabolism
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Brassica Seed ESTs
~ 436,000 seed ESTs
– Various stages of embryo development ~145,000Early seed stages under stress conditions 34 300– Early seed stages under stress conditions ~34,300
– Microspore embryogenesis ~29,000– Germination ~48,900– Dormancy ~33,200– Seed Coat ~121,800– Endosperm ~23 500– Endosperm ~23,500
> All ESTs were deposited in GenBankTTeam: R. Datla, F Georges, J Krochko, A Cutler, E Tsang, G Selvaraj and J Zou
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Proteomic analysis of endosperm with heart stage embryog y
Zou et al
Endosperm in ovules containing p gheart embryos exhibit all phases of endosperm development (Brown et al., 1999)
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P f h BProteome of the B. napusendosperm
• Total proteins identified: 809p
• Chloroplast gene products 17
• Mitochondrial gene products 11Mitochondrial gene products 11
• Genes with no ESTs (matching BNAEN3GH) 149
• Genes with 4 or more EST members 348Genes with 4 or more EST members 348
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Notable entries in the proteome (Endosperm)
Cell wall invertase• Cell wall invertase• Invertase/pectinase inhibitor• cytosolic beta-amylasey y• myo-inositol 1-phosphate synthase• Various components of Golgi vesicle-mediate
t ttransport• Heme transporter• Lipid metabolism enzymes: fatty acyl desaturase; p d e abo s e y es a y acy desa u ase;
acyltransferase• 13 entries correspond to embryo defective genes
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B. napus Seed Coat structure and development
Selvaraj et al
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Proteomic Analysis of Black and Y ll B S d CYellow B. napus Seed Coat
Questions to be addressed:Questions to be addressed:
• What proteins constitute the total proteomes of black and yellow B napus seed coats?of black and yellow B. napus seed coats?
• Is there a discernable difference between the d f bl k (N89 53) dseed coat proteomes of black (N89.53) and
yellow (YN01.429) B. napus seeds?– Do these identifiable differences contribute to the phenotypic differences?
Confidential
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Proteomic Analysis of B. napus Seed Coat Protein ExtractsBlack Seed Coat Yellow Seed Coat
Acidic Basic Acidic Basic
Confidential
Protein identification Protein identification
Protein Comparison
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Brassica embryo/seedBrassica embryo/seed development
Datla et al
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B. napus Embryo Proteins
Globular Heart Torpedo Bent Mature StageskD
250150100
kD
1007550
37
2520
15
10
700 600 40 8 2 #Embryos
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Protein Profiles of Key E b St i B
2000
Embryo Stages in B. napus
1747
12001600
2000
ntifi
edat
ed)
992 1070
428800
1200
Prot
eins
Iden
(Non
-rei
tera
428
0
400
#P
Globular Heart Torpedo Bent Mature
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Proteome of B. napus and Arabidopsis embryos ( l b l t t t )(globular to mature stages)
1032 9391364
60% overlap between identified Arabidopsis and Brassica embryo proteins
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Functional Classification of Embryo Expressed Proteins(Molecular Function)
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Integrated Brassica napus and ArabidopsisEmbryo Metabolic Network
EMB2360
TWN2 EMB2728
EMB2024
EMB1144
PLE2 RML1
EMB1989 EMB2284
EMB2757
EMB1276
EMB1395
Common pathwayBrassicaArabidopsisEmbryo defective
Core reactionNon-core reaction
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Brassica Seed Developmentd M t b liand Metabolism
Example of targets• Large seed size• Uniform seed germination
Red ced coat thickness• Reduced coat thickness
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Crop Genomics for a Healthy Canada A i lt /Pl tAgriculture/Plants
Genome Canada Position paperKeller et al
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Agriculture/Plants(Crop Genomics for a Healthy Canada)(Crop Genomics for a Healthy Canada)
Development of Position Paper:
Science Steering Committee:
Faouzi Bekkaoui (NRC‐PBI), Bruce Coulman (University of Saskatchewan), Adrian Cutler (NRC‐PBI), Raju Datla (NRC‐PBI), Wilf Keller (NRC‐PBI), Isobel Parkin (AAFC‐S k ) J Si h (AA C O )Saskatoon), Jas Singh (AAFC‐Ottawa)
Writer:
Michael Raine
Sponsoring Centre – Genome Prairie:
Jerome Konecsni (CEO), Reno Pontarollo (CSO), Faye Pagdonsolan (Admin Support)
Workshop:p
May 31, 2007, Chateau Laurier Hotel, Ottawa
Approximately 50 participants: Academia, Government, Industry
Facilitator:Facilitator:
Pete Desai (Desai & Desai Inc.)
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Position Paper: Crop GenomicsPosition Paper: Crop Genomicsfor a Healthy Canada
Scope:Scope:
National: cover major Canadian agricultural crops
Key Objectives:• Enhanced crop yield/productivity
• Enhanced health/nutritional value
• Enhanced tolerance to abiotic and biotic stress
• Environmentally friendly, sustainable production
• Di ersification s pport biof el bioprod ct de elopment• Diversification: support biofuel, bioproduct development
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Potential Comp IV Projects – Based on SpeciesProjects Based on Species
• Brassica oilseeds
• Flax
l l il hi k b )• Pulse crops lentils, peas, chickpeas, beans)
• Wheat
• Barley
• OatsOats
• Corn
• Soybean
• Alfalfa (and other forages)
• Potatoes
• Prunus spp (cherries, etc.)
• Blueberries
N l tf d l t ( C li T iti l )• New platform development (e.g. Camelina, Triticale)
Questions:
Genomics resource status?
Overall value of crop?
Critical mass (expertise)?
Receptor capacity (i.e. breeding)? – Breeders must play essential role
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Potential Comp IV Projects B d ThBased on Themes
• Abiotic stress (e.g. heat/drought)
• Biotic stress (e.g. Fusarium)Biotic stress (e.g. Fusarium)
• Plant productivity/yield (e.g. architecture, heterosis)
• Bioproduct development (industrial oils)Bioproduct development (industrial oils)
• Health product development
• Sustainability (e g nutrient use efficiency)• Sustainability (e.g. nutrient use efficiency)
• Comparative genomics
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Canola as an Example: Well Positioned for Genomics Research Projects
• $11B industry
• ~75 public sector scientists (~300 researchers)
• Active private sector (>10 companies)• Active private sector (>10 companies)
• History of genetic research in Canada
• >600,000 ESTs (made in Canada)>600,000 ESTs (made in Canada)
• Microarrys (3 made in Canada)
• TILLING populations (UBC, AAFC)
• International B. rapa Genome Sequencing Initiative
• Arabidopsis data is relevant
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Acknowledgements
• Wilf Keller
For additional information www.brassicagenomics.ca
• Raju Datla, Daoquan Xiang,, Edwin Wang
• Jitao Zou
G l S l j P l A h J h K ll• Gopalan Selvaraj, Paula Ashe, John Kelly
• Andrew Sharpe, Dwayne Hegedus, Kevin Razwodowski, Derek Lydiate, Isobel Parkin
• DOTM and ECTG projects teams
Agri-Food Canada Agroalimentaire CanadaAgriculture and Agriculture et
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