Team:Toulouse/Project/Fungicides

The main objective of SubtiTree is to ensure the destruction of the pathogenic fungi inside the tree. In order to achieve this goal, we built a genetic module to produce three different peptides with antifungal activities.

Originated from plants, these peptides have different target to maximize the lethality on C. platani.

- D4E1 is a synthetic peptide made of 17 amino acids which has been shown to have an antifungal activity by complexing with a sterol present in the conidia’s wall of numerous fungi.

- GAFP-1 (Gastrodia Anti Fungal Protein 1) is a lectin binding to mannose thus inhibiting the growth of a variety of fungi.

- EcAMP-1 (Echinochloa crus-galli Anti Microbial Peptide) consists in 37 amino acids inhibiting hyphae elongation.

More information on this module :

Final constructions: (see Parts to have more details on the intermediate parts) We built different genetic constructions to test each fungicide separately and to test them all together on the same operon where the 3 genes are placed under the control of a constitutive promoter in Bacillus subtilis: Pveg.

EcAMP-1 was already present in the Registry, added by the Utah State 2013 iGEM team (BB_K1162001). We added D4E1 and GAFP-1 to the Registry of Standard Biological Parts (see parts). These new BioBricks were designed in order to be expressed and secreted with Bacillus subtilis.

Secretion :

In order to export the peptides outside the bacteria, the coding sequence of D4E1 and GAFP-1 was flanked on the N-terminal end with a signal peptide (amyE signal peptide) followed by a pro peptide, cleaved during the secretion process.

Chemotaxis

Spreading