Transcriptional regulation of tomato fruit ripening - Molecular Biology

Fruits provide humans with essential nutrients for health benefits, such as vitamins, dietary fiber and minerals. Tomato is the model species for studying fleshy fruit development and ripening.

We study transcription factors RIPENING INHIBITOR (RIN), NON- RIPENING (NOR) and COLORLESS NON-RIPENING (CNR), which are the main factors controlling ripening, together with gaseous hormone ethylene in tomato fruits. Functions of RIN, NOR and CNR have been known already through the study of naturally occurring mutants, but how they work together in a network to control colour, flavour and texture during fruit ripening by regulating many common downstream effector genes, is poorly understood. We aim to unravel the mechanisms of combinational action of tomato transcription factors on the expression of relevant ripening-related genes through:

  • Studying the effector genes of combining different gene dosages of the major regulators on fruit ripening using heterozygous progeny of different crossed wild type and mutant parents (using natural mutations as well as our own generated CRISPR/CAS9-mutants)
  • Studying downstream gene promoter architecture using targeted deletion with CRISPR/CAS9

We are experienced in editing genome by using CRISPR/CAS9 and are continuing to develop new strategies in mutagenesis. Null mutants in genes encoding major transcription factors have been obtained and promoter mutagenesis experiments by multiple guide-RNAs are ongoing.

Used skills

  • Bioinformatics tools
  • Molecular cloning and sequencing
  • Protein (-DNA) interaction studies with yeast 1, 2- or 3- hybrid assays
  • Gene expression analysis with qRT-PCR
  • CRISPR/CAS9 mutagenesis and mutant characterization and phenotyping
  • Plant transformation and tissue culture

Requirements

  • Good theoretical and practical basis in (plant) molecular biology

 

Phenotype of T0 mutants generated by using CRISPR/CAS9. rin single mutant (left), nor and rin double mutant (middle) and wild type Moneyberg (right).
Phenotype of T0 mutants generated by using CRISPR/CAS9. rin single mutant (left), nor and rin double mutant (middle) and wild type Moneyberg (right).