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Experiment: Impact of Type III effectors on plant defense responses
Experiment Description
NASCArrays Experiment Reference Number: NASCARRAYS-59
Our interest lies in how plants respond to bacterial pathogens. Over the past three years we have identified and documented reproducible, landmark biochemical and molecular events following the challenge of Arabidopsis with the phytopathogenic enterobacteria P. syringae. Significantly, our studies revealed 60% of cDNA-AFLP differentials not present on the 8,200 feature GeneChips and 20% absent from public EST databases (de Torres in press). We now seek to exploit this background using carefully defined time-points to analyse global changes in the Arabidopsis transcriptome using challenges selected to define gene targets implicated in (i) expression of basal immunity (ii) the establishment of successful parasitism (resistance) by a virulent pathogen (host). The results will provide a rationale for future functional assays of the identified pathways using transgenic knockouts and mutant analyses. Additionally, data will provide underpinning support for comparative proteomics of the defense response currently in progress with GARNet support using the same experimental parameters (BBSRC 32/P14635).
We propose the following treatments:
(i) Mock vs. DC3000hrpA @ 60 min: 60 minutes is subsequent to host immediate-early stress responses and will catalogue the innate responses induced by pathogen associated molecular patterns. The hrpA lesion will ensure no type III effectors influence transcriptional responses. Gene products induced at this time are predicted to potentiate latter host responses (2 treatments X 3 biological replicates = 6 chips).
(ii) Mock vs. DC3000 vs. DC3000hrpA vs. DC3000::avrRpm1 @ 4 hours: A key time point previously defined where no macroscopic symptoms are visible but significant differences exist between compatible and incompatible interactions at the molecular and physiological levels. These treatments will serve to define the earliest genes induced by the complement of DC3000 type III effector and will specifically define genes/pathways suppressed by virulence factors in addition to those implicated in orchestration of the hypersensitive cell death. We will also include an incompatible interaction on a transgenic line expressing an RPM1 interacting protein, which fails to mount an HR but exhibits hyper-resistance. We predict this challenge will separate the resistance response (pathogen restriction) from that associated with hypersensitive cell death (5 treatments X 3 biological replicates = 15 chips).
(iii) Mock vs. DC3000 vs. DC3000hrpA @ 14 hours: At 10 h before phenotypes are apparent in the DC3000 background, Type III effector delivery is well advanced and impact on host transcription maximal (3 treatments X 3 biological replicates = 9 chips).
About the ExperimenterName: | Dr Marta de Torres Zabala |
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Head of Lab Name: | Dr Murray Grant |
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Lab:
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Institute:
| Imperial College at Wy |
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Address: | Department of Agricultural Sciences Imperial College at Wye Wye
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Postcode:
| TN25 5AH |
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Country:
| UK |
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| Telephone Number:
| 02075942883 |
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Fax Number:
| 02075942640 |
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All of the data available in this website/database is free, and you
are free to do whatever you please with it. If you intend to publish
work based on any of this data, please acknowledge us, contact the
experimenter above, and either acknowledge them or use them as
co-authors in the work.
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| About this ExperimentExperiment Type:
| time_series_design; pathogenicity_design; compound_treatment_design |
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Number of Slides: | 27 |
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| Experimental Parameters:
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Parameter | compound_based_treatment |
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parameter | age |
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parameter | infect |
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Quality Control Measures Taken:
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no-plants-pooled | 4 leaves/plant; 18 plants/time point |
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References:
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| Other Information:
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Slides in this Experiment
Hybridisation Set: de Torres Zabala Rep1
Slide: A-1-Torres-MgCl2_2hr_Rep1 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
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treatment | "Leaves were challenged with 10 mM MgCl2. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syring
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment." |
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Other Information:
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Protocols for BioSource 1 |
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Slide: A-4-Torres-Hrp_2hr_Rep1 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
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treatment | Leaves were challenged with the hrpA mutant of bacterium Pseudomonas syringae. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe containing the bacterium at a concentration of 7 X 107 colony forming units/ml in 10 mM MgCl2.
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment. |
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Other Information:
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Protocols for BioSource 1 |
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Slide: A-7-Torres-MgCl2_4hr_Rep1 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | "Leaves were challenged with 10 mM MgCl2. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syring
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment." |
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Other Information:
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Protocols for BioSource 1 |
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Slide: A-10-Torres-Hrp_4hr_Rep1 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with the hrpA mutant of bacterium Pseudomonas syringae. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe containing the bacterium at a concentration of 7 X 107 colony forming units/ml in 10 mM MgCl2.
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment. |
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Other Information:
| |
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Protocols for BioSource 1 |
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Slide: A-13-Torres-DC3000_4hr_Rep1 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
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treatment | Leaves were challenged with Pseudomonas syringae pv. tomato DC3000 (susceptible on Col-5). Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe at a bacterium concentration of 1 X 108 colony forming units/ml.
At least 4 leaves from 18 plants were used in this treatment, and leaves were harvested 4 hours later. |
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Other Information:
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Protocols for BioSource 1 |
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Slide: A-16-Torres-AvrRpm1_4hr_Rep1 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
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treatment | Leaves were challenged with Pseudomonas syringae pv.tomato DC3000 carrying the avrRpm1 avirulence gene (resistant to Col-5). Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe at a bacterium concentration of 1 X 108 colony forming units/ml.
At least 4 leaves from 18 plants were used in this treatment, and leaves were harvested 4 hours later. |
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Other Information:
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Protocols for BioSource 1 |
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Slide: A-22-Torres-MgCl2_12hr_Rep1 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with 10 mM MgCl2. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syring
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment. |
---|
Other Information:
| |
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Protocols for BioSource 1 |
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Slide: A-25-Torres-DC3000_12hr_Rep1 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with Pseudomonas syringae pv. tomato DC3000 (susceptible on Col-5). Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe at a bacterium concentration of 1 X 108 colony forming units/ml.
At least 4 leaves from 18 plants were used in this treatment, and leaves were harvested 12 hours later. |
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Other Information:
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Protocols for BioSource 1 |
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Slide: A-28-Torres-Hrp_12hr_Rep1 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
---|
treatment | Leaves were challenged with the hrpA mutant of bacterium Pseudomonas syringae. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe containing the bacterium at a concentration of 7 X 107 colony forming units/ml in 10 mM MgCl2.
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment. |
---|
Other Information:
| |
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|
Protocols for BioSource 1 |
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Hybridisation Set: de Torres Zabala Rep2
Slide: A-2-Torres-MgCl2_2hr_Rep2 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | "Leaves were challenged with 10 mM MgCl2. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syring
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment." |
---|
Other Information:
| |
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Protocols for BioSource 1 |
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Slide: A-5-Torres-Hrp_2hr_Rep2 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with the hrpA mutant of bacterium Pseudomonas syringae. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe containing the bacterium at a concentration of 7 X 107 colony forming units/ml in 10 mM MgCl2.
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment. |
---|
Other Information:
| |
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Protocols for BioSource 1 |
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Slide: A-8-Torres-MgCl2_4hr_Rep2 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | "Leaves were challenged with 10 mM MgCl2. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syring
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment." |
---|
Other Information:
| |
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|
Protocols for BioSource 1 |
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Slide: A-11-Torres-Hrp_4hr_Rep2 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with the hrpA mutant of bacterium Pseudomonas syringae. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe containing the bacterium at a concentration of 7 X 107 colony forming units/ml in 10 mM MgCl2.
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment. |
---|
Other Information:
| |
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|
Protocols for BioSource 1 |
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Slide: A-14-Torres-DC3000_4hr_Rep2 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with Pseudomonas syringae pv. tomato DC3000 (susceptible on Col-5). Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe at a bacterium concentration of 1 X 108 colony forming units/ml.
At least 4 leaves from 18 plants were used in this treatment, and leaves were harvested 4 hours later. |
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Other Information:
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|
Protocols for BioSource 1 |
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Slide: A-17-Torres-AvrRpm1_4hr_Rep2 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with Pseudomonas syringae pv.tomato DC3000 carrying the avrRpm1 avirulence gene (resistant to Col-5). Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe at a bacterium concentration of 1 X 108 colony forming units/ml.
At least 4 leaves from 18 plants were used in this treatment, and leaves were harvested 4 hours later. |
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Other Information:
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|
Protocols for BioSource 1 |
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Slide: A-23-Torres-MgCl2_12hr_Rep2 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with 10 mM MgCl2. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syring
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment. |
---|
Other Information:
| |
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|
Protocols for BioSource 1 |
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Slide: A-26-Torres-DC3000_12hr_Rep2 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with Pseudomonas syringae pv. tomato DC3000 (susceptible on Col-5). Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe at a bacterium concentration of 1 X 108 colony forming units/ml.
At least 4 leaves from 18 plants were used in this treatment, and leaves were harvested 12 hours later. |
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Other Information:
| |
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|
Protocols for BioSource 1 |
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Slide: A-29-Torres-Hrp_12hr_Rep2 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with the hrpA mutant of bacterium Pseudomonas syringae. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe containing the bacterium at a concentration of 7 X 107 colony forming units/ml in 10 mM MgCl2.
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment. |
---|
Other Information:
| |
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|
Protocols for BioSource 1 |
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Hybridisation Set: de Torres Zabala Rep3
Slide: A-3-Torres-MgCl2_2hr_Rep3 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | "Leaves were challenged with 10 mM MgCl2. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syring
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment." |
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Other Information:
| |
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Protocols for BioSource 1 |
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Slide: A-6-Torres-Hrp_2hr_Rep3 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with the hrpA mutant of bacterium Pseudomonas syringae. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe containing the bacterium at a concentration of 7 X 107 colony forming units/ml in 10 mM MgCl2.
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment (replicate #3). |
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Other Information:
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Protocols for BioSource 1 |
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Slide: A-9-Torres-MgCl2_4hr_Rep3 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | "Leaves were challenged with 10 mM MgCl2. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syring
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment." |
---|
Other Information:
| |
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|
Protocols for BioSource 1 |
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Slide: A-12-Torres-Hrp_4hr_Rep3 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with the hrpA mutant of bacterium Pseudomonas syringae. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe containing the bacterium at a concentration of 7 X 107 colony forming units/ml in 10 mM MgCl2.
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment. |
---|
Other Information:
| |
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|
Protocols for BioSource 1 |
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Slide: A-15-Torres-DC3000_4hr_Rep3 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with Pseudomonas syringae pv. tomato DC3000 (susceptible on Col-5). Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe at a bacterium concentration of 1 X 108 colony forming units/ml.
At least 4 leaves from 18 plants were used in this treatment, and leaves were harvested 4 hours later. |
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Other Information:
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|
Protocols for BioSource 1 |
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Slide: A-18-Torres-AvrRpm1_4hr_Rep3 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with Pseudomonas syringae pv.tomato DC3000 carrying the avrRpm1 avirulence gene (resistant to Col-5). Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe at a bacterium concentration of 1 X 108 colony forming units/ml.
At least 4 leaves from 18 plants were used in this treatment, and leaves were harvested 4 hours later. |
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Other Information:
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|
Protocols for BioSource 1 |
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Slide: A-24-Torres-MgCl2_12hr_Rep3 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with 10 mM MgCl2. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syring
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment. |
---|
Other Information:
| |
---|
|
Protocols for BioSource 1 |
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Slide: A-27-Torres-DC3000_12hr_Rep3 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with Pseudomonas syringae pv. tomato DC3000 (susceptible on Col-5). Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe at a bacterium concentration of 1 X 108 colony forming units/ml.
At least 4 leaves from 18 plants were used in this treatment, and leaves were harvested 12 hours later. |
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Other Information:
| |
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|
Protocols for BioSource 1 |
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Slide: A-30-Torres-Hrp_12hr_Rep3 | | |
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Tissue:
| Whole leaf |
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Diseased:
| Normal |
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in vitro Treatment:
| |
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treatment | Leaves were challenged with the hrpA mutant of bacterium Pseudomonas syringae. Both sides of a leaf were infiltrated on their abaxial surface with a needleless 1ml syringe containing the bacterium at a concentration of 7 X 107 colony forming units/ml in 10 mM MgCl2.
This treatment is equivalent to a mock challenge to mimic local wounding responses.
At least 4 leaves from 18 plants were used in this treatment. |
---|
Other Information:
| |
---|
|
Protocols for BioSource 1 |
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Problems? Comments? Suggestions? Contact the Affymetrix Team