RNAi Expression in Cotton for Control of Herbivorous Insects.

ABSTRACT

Cultivated cotton (Gossypium hirsutum) is heavily attacked by various species of insects worldwide and breeding of new varieties resistant to pests is still a hard battle to win. RNAi technology is an important reverse genetics tool to induce gene silencing in eukaryotic organisms and produce phenotypic modifications. In cotton, RNAi was applied to investigate gene function and enhance resistance to insects and pathogens. Different methods and techniques can be used to synthetize double stranded RNA (dsRNA) into plant cells. The Agrobacterium-mediated transformation is a common method to introduce RNAi binary plasmids into cotton genome and obtain stable transgenics plants. This methodology includes the coculture of cotton tissues with Agrobacterium cultures, selection of transgenic cells and induction of somatic embryogenesis to finally obtain transgenic plants after a relatively long period of time. The transient synthesis of dsRNA mediated by virus-induced gene silencing (VIGS) in cotton is an alternative to anticipate the silencing effect of a specific RNA sequence, prior to the development of a stable transgenic plant. VIGS vectors are incorporated into the plant by agroinfiltration technique. During VIGS replication inside plant cells, synthetized dsRNA allows the study on specific heterologous gene expression including the phenotypic effect on herbivorous target pests, thus facilitating a rapid evaluation of dsRNA expressed in cotton plants against individual insect target genes. Here we describe the complementation of these two techniques to evaluate RNAi-based cotton plant protection against insect pests.