Development of In Vitro Root Culture and miRNAs Analysis for Secondary Metabolites of Native Plants from the Mexican Bajio ^†

Mexico is a megadiverse country, with a high quantity of unique plant species with different uses and applications, such as bactericidal, fungicidal, insecticidal, and, recently, nutrimental. The content of phytochemicals, and the impact of them on animal and human health, has made them a target for biotechnological improvement. In the region Bajio in Mexico, several plants that are associated with ecological, medical and industrial potential have been identified, but also those associated with the traditions. The work in this project includes the development of systems for the culture for the production of secondary metabolites (in vitro root tissue culture) and the miRNA expression analysis, in order to find the molecules that are associated with metabolites production. In this study, we include the following two plants: marigold (Tagetes erecta), in which genes associated to lutein production had been identified in flower development; systems for cell culture and plant transformation have been developed, but no systems for in vitro root culture. Up until now, there are not studies related to miRNA expression and association to these molecules to secondary metabolites. In Heliopsis longipes, several methodologies had been developed for the isolation of afinnin and its uses in agriculture, medicine, and, recently, as analgesic activities in some other metabolites. First, a root tissue culture was established for both of the plants (marigold and Heliopsis), using a combination of auxins (2,4-D, IAA, IBA) in a kinetic assay, as the base for manipulation; differences in the root architecture were determined mainly in the time of production and root architecture. In the molecular analysis, four miRNAs were found to be differentially expressed, and associated to secondary metabolites production (miR146, miR164, miR168, miR171). The reordering of miRNAs synthesis and the targets was analyzed, and is associated with the secondary metabolites production, in order to establish a system for the in vitro induction of metabolites.