Projets - V (Victor) Aprilyanto

Investigating molecular regulation of fruit ripening and shelf life

CRISPR/Cas technology opens huge possibilities in studying the role of genes in regulatory networks, including fruit ripening and shelf life. By applying CRISPR/Cas-targeted gene knockout, one can study how fruit ripening commences through series of interactions among transcription factors and their immediate target genes. 

Three transcription factors (TFs) called 'master regulators', namely MADS-RIN, SPL-CNR and NAC-NOR, regulate tomato fruit ripening by activating or repressing downstream target genes. In addition to NAC-NOR, several other NAC TFs (NOR-like1, NAC4, NAC1, and NAM1) are known to affect fruit development and ripening. There is also GRAS38, a member of the GRAS family TF, which was recently known to affect fruit firmness. Several studies have addressed the putative function of these genes by RNA interference or virus-induced gene silencing techniques. However, due to RNAi and VIGS's specificity problem, it is difficult to tell whether the observed result resulted from complete or partial target gene inactivation or off-target results. My first aim is to re-evaluate the NAC and GRAS38 TFs functions by producing knockouts using the CRISPR/Cas system. This study will include creating single- and multiple-gene knockout mutants. After that, I want to study the interaction among TF proteins to gain insights into formation of protein complexes for activation or repression activity. 

Tomato will be used as a model plant throughout this study. The knowledge of fruit ripening mechanisms and their immediate consequences for fruit shelf life could be directly applied to one of the most consumed fruit in the world. 

If you like to work on these projects, you can send me an email . 

Used skills: 

• Basic molecular biology techniques (PCR, cloning, restriction enzyme etc.) 
• Basic bioinformatics
• Golden Gate Assembly 
• Plant transformation and tissue culture
• Mutant analysis & plant phenotyping