0 avis
Developing gene editing strategies in Tomato as a method to translate new genetic resistance to viruses.
Archive ouverte
Edité par CCSD -
International audience. A major goal of plant breeding is to be able to translate genetic resistances to viruses across species and in the genetic background of choice. In the case of susceptibility factors, it is thought that resistance to viruses can be achieved by genome editing following the inactivation (knock-out) of Susceptibility genes. Here, we will present results on three candidate genes whose role on resistance to viruses have been previously demonstrated in Arabidopsis. We present our strategy to develop gene inactivation, in two different tomato genetic background. This allow us to try to i) develop resistances to pepino mosaic virus and potexvirus, which are a threat to tomato crops worldwide but also ii) to aim at enhancing the durability of resistance to potyvirus driven by an eIF4E weak allele. We describe a general procedure using CRISPR-Cas9 strategy to achieve gene knockout in tomato following agrobacterium mediated plant transformation. The targeting a genomic coding sequence of a gene of interest allows to generate indels following Double Strand Breaks (DSB). We will therefore present the different steps from designing the sequence guide RNA (sgRNA), cloning the binary vector that will express in planta both sgRNA and the Cas9 nuclease, to agrobacterium-mediated plant transformation. Following regeneration, we will present the characterization of heterozygous and homozygous gene mutated alleles. Because our aim is to contribute to plant breeding, we assess both the potential effect on resistance to viruses, but also how knocking-out Susceptibility gene may affect tomato plant development. Indeed, we report several plant and fruit abnormal phenotypes associated with those genes’ knock-out, such as a reduction in plant size or abnormal fruit shape and content. This makes of these line interesting tools to investigate developmental biology in Tomato, but together with preliminary resistance assays, suggests that overall, knocking-out those genes is unlikely to constitute strong sources of resistance for Tomato breeding.
Consulter en ligne
Suggestions
Du même auteur
