Fountain of Youth - Molecular Biology

Thesis subjects Discovering the Fountain of Youth: a chemical genetics approach to identify enhancers of Arabidopsis somatic embryogenesis

Description

Plants are perhaps the most developmentally plastic of all organisms. Not only do they continually differentiate new organs from the stem cell niche throughout their lifespan, but they are also capable of regenerating new cell types and organs after wounding or from explants in tissue culture. One example of this plasticity is somatic embryogenesis (SE), where embryos form on vegetative organs in the absence of fertilization. SE  is most commonly induced by exposing explants to stress conditions and/or exogenous growth regulators such as the synthetic auxin 2,4-dichlorophenoxyacetic acid (2,4-D). The molecular and cellular basis for SE, as well as other plant regeneration processes, is poorly understood. We addressed this knowledge gap by using a chemical genomics approach to identify compounds that enhance SE in arabidopsis seedlings. Two compounds were identified that induce SE in ~50-70% of the seedlings compared to ~20% in the control. The aim of this project is to identify the target proteins and downstream pathways affected by these two compounds. A number of approaches are being used including analysis of auxin mutants, analysis of GFP reporter lines, analysis of gene expression changes after compound treatment by microarray analysis, and immunopreciptation of compound-target protein complexes.

Used skills

  • Genetic analysis of arabidopsis mutants
  • Analysis and verification of microarray data (bioinformatics, DNA and RNA isolation, qRT-PCR)
  • Development of arabidopsis GFP reporter- and overexpression lines (PCR, Gateway Cloning)
  • Transformation of Arabidopsis
  • Analysis of GFP reporter lines using Confocal Laser Scanning Micrsoscopy
  • Protein biochemistry (protein isolation, immunopreciptation, Western blotting)
  • Tissue culture / somatic embryo culture

Requirements

  • Theoretical and practical basis in molecular biology
  • Affinity for plant tissue culture