S19 - Session O3 - A HIGS approach targeting the DCL1, Cyp51 and CHS genes of the pathogen to control Colletotrichum acutatum infection of strawberry

Authors: José-Javier Higuera-Sobrino, Victoria Súnico, Lucía Cervantes-Cárdenas, Rosario Blanco-Portales, Enriqueta Moyano, Juan Muñoz-Blanco, José Luis Caballero Repullo *

Among the berries, the strawberry ( Fragaria x ananassa ) represents one of the most valuable cultivated species worldwide. However, this crop suffers significant losses due to attack by many fungal pathogens including Colletotrichum acutatum , which devalues the fruit at the harvest and post-harvest stages. To date, due to the lack of alternatives that are really effective, chemical and fungicides are being used to control C. acutatum infection despite the fact that these procedures are not environmentally friendly enough and are also risky for the Human health. New biotechnological approaches, such as h ost i nduced g ene s ilencing (HIGS) applied to this species, emerge as a valuable alternative to control pathogen infection while reducing the use of chemicals and therefore the impact environmental. In this study, using the Colletotrichum databases currently available, several hairpin-based gene cassettes which target five important pathogen genes, i.e. the dcl-1 and dcl-2 , the cytochrome p450-51A and 51B ( cyp51A/cyp51B) ) and the chitin synthase class V ( chsV ) have been designed and their DNA sequences either single or combined have been obtained by synthetic biology and cloned in gateway derivative T-vectors under the control of the 35S promoter. Putative HIGS lines of strawberry ( F x ananassa ) cv A1 (Viveros California) have been obtained by agrobacterium transformation and kanamycin-based selection. Positive HIGS lines have been molecular characterized by PCR analysis. The efficacy of dsRNAs stably expressed in the HIGS plants is being preliminary tested by inoculating detached leaves from in vitro elongated shoots of strawberry lines. This study is part of the Med-Berry project funded by PRIMA (UE) and AEI (Spain) aimed to develop new strategies to protect strawberry crop in Mediterranean countries.