Somatic embryogenesis - Molecular Biology

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.

Description

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). Research in our lab has shown that 2,4-D-induced SE from germinating Arabidopsis thaliana (arabidopsis) seeds intersects with two additional hormone pathways, abscisic acid (ABA) and brassinosteroids (BR). Molecular-genetic analysis of arabidopsis mutants revealed a role for ABA in promoting SE at three different levels: ABA biosynthesis, ABA receptor complex signalling and ABA-mediated transcription. A chemical screening approach identified a small molecule, brassinomide (BRM), that is a powerful enhancer of SE from germinating seed. BRM inhibits brassinosteroid biosynthesis, which results in auxin- and ABA hypersensitivity during SE. In this project, a range of approaches and techniques will be used to determine how the ABA, brassinosteroid and 2,4-D pathways control SE, and where these pathways intersect.

Used skills

  • CRISPR mutagenesis in arabidopsis (cloning constructs and identification and analysis of mutants)
  • Floral dip transformation
  • Genetic analysis of arabidopsis mutants
  • Transcriptome analysis (RNA-seq, bioinformatics)
  • RNA isolation and qRT-PCR
  • Development of arabidopsis GFP reporter- and overexpression lines (PCR, Gateway cloning)
  • Analysis of GUS (histochemical assays) and GFP reporter lines (confocal laser scanning microscopy)
  • Tissue culture (somatic embryo culture)

Requirements

  • Good theoretical and practical basis in (plant) molecular biology
  • Affinity for plant tissue culture

Students

Would you like to work on these projects? Then send me an e-mail as soon as possible!