Progress 01/01/04 to 12/31/04
Outputs
The long-term goal of this project is to identify regulatory genes that modulate the expression of multiple stress-response pathways. Toward this goal, we are using an activation-tagging approach to identify gain-of-function and loss-of-function mutants that influence stress-response gene expression. This strategy will allow us to dissect the signaling pathways that control the basic developmental and stress-responses for two genes known to respond to multiple environmental stresses: le25 and LapA. Both le25 and LapA respond to environmental stresses that are especially relevant to agriculture in the arid/semiarid environment of the southwestern US including water-deficit, salinity, and/or pest and pathogen attack. We have generated reporter lines that contain either the le25 promoter or the LapA promoter fused to LUCIFERASE. These lines will be useful for the experiments in this project, as well as additional studies on stress-regulated gene expression. We have also
constructed a T-DNA vector containing promoters that will activate neighboring gene expression (activation tagging). In the past year, we have transformed the activation-tagging construct into parental reporter plants. Approximately 20,000 transgenic plants have been generated with this construct. When inserted nearby a chromosomal gene, the activation-tagging construct will cause the mis-expression of that gene. Transgenic plants have been screened for the inappropriate expression of the reporter gene in the absence of stress, which would indicate the identification of a regulator of the stress-response gene. So far, transgenic plants that inappropriately express the reporter gene have not been identified, however a number of plants that display interesting morphological phenotypes have been identified. The insertion site has been determined for a number of these lines. Additional screening is ongoing.
Impacts
In order to fully understand a plant's response to environmental stresses, we need to identify genes that regulate individual and multiple stress-response pathways. Progress in this direction could eventually allow the engineering of crop plants such that they are better able to survive the harsh environmental conditions found in the field.
Publications
- No publications reported this period
|
Progress 08/15/02 to 08/14/04
Outputs
The long-term goal of this project was to identify regulatory genes that modulate the expression of multiple stress-response pathways. We used an activation-tagging approach to identify gain-of-function and loss-of-function mutants that influence stress-response gene expression. We generated transgenic Arabidopsis plants containing the LUCIFERASE reporter gene under control of stress-responsive promoters. We constructed a T-DNA vector containing promoters that will activate neighboring gene expression (activation tagging). When inserted nearby a chromosomal gene, the activation-tagging construct is expected to cause the mis-expression of that gene. We then transformed the activation-tagging construct into parental reporter plants and screened the progeny to identify lines that showed inappropriate expression of the reporter gene in the absence of stress, which would indicate the identification of a regulator of the stress-response gene. Approximately 40,000 transgenic
plants were screened. Unfortunately, we did not identify any insertions that consistently mis-expressed the reporter gene. However we did identify approximately 50 lines that showed interesting morphological phenotypes. The site of insertion was characterized for about 20 of these lines, and analysis of the phenotypes and mis-expressed genes is ongoing.
Impacts
The purpose of this research program is to identify genes that individual and multiple stress response pathways in plants. This will be accomplished by isolating dominant mutations that cause the de-regulation of stress-responsive genes. This project permits the identification and characterization of new genes involved in stress responses.
Publications
- No publications reported this period
|
Progress 01/01/03 to 12/31/03
Outputs
The long-term goal of this project is to identify regulatory genes that modulate the expression of multiple stress-response pathways. Toward this goal, we are using an activation-tagging approach to identify gain-of-function and loss-of-function mutants that influence stress-response gene expression. This strategy will allow us to dissect the signaling pathways that control the basic developmental and stress-responses for two genes known to respond to multiple environmental stresses: le25 and LapA. Both le25 and LapA respond to environmental stresses that are especially relevant to agriculture in the arid/semiarid environment of the southwestern US including water-deficit, salinity, and/or pest and pathogen attack. We have generated reporter lines that contain either the le25 promoter or the LapA promoter fused to LUCIFERASE. These lines will be useful for the experiments in this project, as well as additional studies on stress-regulated gene expression. We have also
constructed a T-DNA vector containing promoters that will activate neighboring gene expression (activation tagging). To determine the feasibility of generating the necessary number of transgenic plants, we initiated a pilot activation-tagging experiment in a background containing the GUS reporter gene driven by the LATERAL ORGAN BOUNDARIES (LOB) promoter. The LOB promoter drives expression at leaf bases and is regulated by hypoxic stress and ABA. We performed transformations estimated to give rise to 10,000 activation-tagged transgenic plants. Approximately 5,000 transgenic plants have been isolated and screened for GUS reporter-gene expression. We are looking for plants that show ectopic GUS activity, which would indicate the activation of an upstream regulator of LOB. Among the 5,000 lines identified, we have documented approximately 20 plants that have interesting morphological defects, but so far we have not identified plants that display ectopic GUS activity. We are currently
introducing the activation-tagging construct into le25-LUC and LapA-LUC backgrounds, and transgenic plants will be screened for LUC activity in the absence of stress.
Impacts
In order to fully understand a plant's response to environmental stresses, we need to identify genes that regulate individual and multiple stress-response pathways. Progress in this direction could eventually allow the engineering of crop plants such that they are better able to survive the harsh environmental conditions found in the field.
Publications
- No publications reported this period
|
|