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Genetical genomics of Puccinia graminis TTKS response: A systems biology approach to rapid development of durable resistance for barley and wheat. Roger Wise USDA-ARS / Iowa State University http://wiselab.org/ http://plexdb.org/ http://barleygenome.org/. Gene Expression and QTLs. - PowerPoint PPT Presentation
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Roger WiseRoger Wise
USDA-ARS / Iowa State UniversityUSDA-ARS / Iowa State Universityhttp://wiselab.org/ http://plexdb.org/
http://barleygenome.org/
Genetical genomics of Genetical genomics of Puccinia graminisPuccinia graminis TTKS TTKS
response: response: A systems biology approach A systems biology approach
to rapid development of to rapid development of durable resistance for barley durable resistance for barley
and wheatand wheat
Gene Expression and QTLsGene Expression and QTLs
The treatment of transcript The treatment of transcript accumulation as a Quantitative Trait is accumulation as a Quantitative Trait is called eQTL mapping. Loci that regulate called eQTL mapping. Loci that regulate expression of genes or networks can be:expression of genes or networks can be:
-cis-cis-acting QTL-acting QTL-trans-trans-acting QTL-acting QTL
RequirementsRequirements
• Experimental population that harbors genetic Experimental population that harbors genetic
variation for resistance to the pathogen variation for resistance to the pathogen
[QSM DH (Q21861 x SM89010)][QSM DH (Q21861 x SM89010)]
• Global gene expression profiling platform Global gene expression profiling platform
(Affymetrix Barley1 GeneChip)(Affymetrix Barley1 GeneChip)
• High-throughput genotyping platform High-throughput genotyping platform
(QSM (QSM SFPs detected by Barley1 GeneChip)SFPs detected by Barley1 GeneChip)
• Database Database (BarleyBase/PLEXdb)(BarleyBase/PLEXdb)
Scoring of TTKS on QSM DH mapping Scoring of TTKS on QSM DH mapping populationpopulation
0;0; 1212 2121 33--210;210; 33++
Brian SteffensonBrian Steffenson
Barley1 GeneChipBarley1 GeneChip 22,840 Probe Sets 22,840 Probe Sets
((22,792 + controls22,792 + controls))
Perfect Match 25-merMismatch 25-mer600 bp5´ 3´AAAAAAAAAAAATGTAGGene SequenceClose Close et al.et al. 2004, Pl Physiology 134: 960-968 2004, Pl Physiology 134: 960-968
Single feature Polymorphisms (Steptoe and Morex)
Raw Data:Steptoe
Morex
Robbie WaughRobbie Waugh
22,000 Genes X11 Oligos X 25nt = 6,325,000 bases
[ >4,000 SFPs ]
Contig3660_atContig3660_at
Expression Level Polymorphism
Matt MoscouMatt Moscou
Experimental Design
Each flat contains up to 98 cones of (6) seedlings randomly arranged among reps and inoc & mock-inoc (Harvested 24 hours after
inoculation).
RNA from each cone ofseedlings is hybridizedto its own Barley1GeneChip.
Diversity Set (12 lines)
Red Trays- Inoculated
Blue Trays- Mock-inoculated
Dew Chamber
[79 DH lines + parents] x 2 inoc. treatments x 2 reps = 340 randomized samples
QSM DH population (79 lines)
12 diversity lines x 2 inoc. treatments x 6 reps [= 144 randomized samples]
Data AnalysisData Analysis
• Identify subset of genes that show Identify subset of genes that show differential expression between TTKS-differential expression between TTKS-inoculated and mock-inoculated leaves. inoculated and mock-inoculated leaves.
• Map the eQTL that regulate the expression Map the eQTL that regulate the expression of these genes.of these genes.
• Identify regulators by hierarchical analyses.Identify regulators by hierarchical analyses.
• A locus that regulates one or more A locus that regulates one or more significant disease resistance networks is an significant disease resistance networks is an obvious target for immediate use in obvious target for immediate use in breeding. breeding.
ExpectationsExpectations
1. 4,000-8,000 SFP gene markers positioned on the barley population segregating for TTKS resistance; enables QTL mapping of TTKS resistance.
2. Marker-delineated positions of the eQTL that regulate networks of disease response genes; breeders can incorporate into their germplasm.
3. Elucidation of gene expression networks responsive to TTKS infection; provides biochemical targets for further analysis.
4. Genes that possess transcriptional activation elements that allow them to respond more aggressively to TTKS infection; provides key regulators for disease response.
What do we do with all that data?What do we do with all that data?(65 Mb/chip = > 30 GigaBytes)(65 Mb/chip = > 30 GigaBytes)
http://www.plexdb.org
1000 Barley1 Hybs Uploaded1000 Barley1 Hybs Uploaded
USDA-ARS/ISUUSDA-ARS/ISURico Caldo Rico Caldo
Matt MoscouMatt MoscouGreg FuerstGreg FuerstNick LauterNick Lauter
Jose RodriguezJose RodriguezYan MengYan Meng
Pingsha Hu Pingsha Hu Liu XiLiu Xi
Dennis Halterman Dennis Halterman Karin WernerKarin Werner
Brent KronmillerBrent Kronmiller
BarleyBase/PLEXdbBarleyBase/PLEXdbJulie DickersonJulie DickersonEthy CannonEthy Cannon
Sudhansu DashSudhansu DashLu HongLu Hong
Lishuang ShenLishuang ShenDan NettletonDan Nettleton
USDA-ARS/UMNUSDA-ARS/UMN Yue JinYue Jin
Sam GaleSam GaleLes Szabo Les Szabo
Univ. of MinnesotaUniv. of MinnesotaBrian SteffensonBrian SteffensonStephanie DahlStephanie Dahl
Knowledge For Tomorrow’s Solutions
http://barleygenome.org
http://barleycap.org