Evaluation of the efficiency of virus-induced gene silencing in different Solanaceae species

• Grant number: 19/27045-0
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: February 01, 2020
• End date: October 31, 2022
• Field of knowledge: Biological Sciences - Genetics - Plant Genetics
• Principal Investigator: Paulo José Pereira Lima Teixeira
• Grantee: Ana Luisa Ramos de Oliveira
• Host Institution: Escola Superior de Agricultura Luiz de Queiroz (ESALQ). Universidade de São Paulo (USP). Piracicaba , SP, Brazil
• Associated research grant: 18/24432-0 - Functional characterization of effector proteins from cacao pathogens and search for their cognate immune receptors, AP.JP

Abstract

Genetic manipulation tools are essential for the study of gene function in anyorganism. In plants, multiple strategies can be adopted for gene inactivation, includingchemical mutagenesis, insertion of T-DNAs or transposons and site-directed mutagenesis by CRISPR/Cas9, ZFNs or TALENs. However, the generation of large-scale mutant populations and the identification of mutants of interest is a laborious and time-consuming process. RNAi silencing using stable transformation is another strategy used in the study of gene function, but this method is considerably slow because it depends on genetic transformation and the advancement of generations. On the other hand, virus-induced gene silencing(VIGS) is a widely used technique for silencing genes of interest in multiple species.mainly due to its practicality and the fact that it does not depend on stable transformation. The VIGS technique uses the natural plant defense system against viruses to promote the silencing of genes of interest. This technique has already been successfully used in genetic analyses in a number of plant species, including tobacco, tomatoes, pepper, rose, beans, soybeans, among others. Thus, this project aims to evaluate the efficiency of the VIGS technique in a collection of Solanaceae aiming to establish the gene silencing method for future genetic analysis. Vectors containing a phytoene gene fragment (PDS, phytoene desaturase) will be constructed and used to silence the carotenoid synthesis pathway in the target plants. The PDS enzyme converts the colorless phytoene molecule into carotenoids, which absorb light in the spectrum of the visible region. Loss of PDS gene function results in white leaves due to the loss of photobleaching protection, allowing for easy detection of silencing events. The establishment of the VIGS technique in multiple solanaceous species will allow future functional analyses of specific genes related to plant resistance against pathogens. (AU)