Analysis of callus development and transient expression in Tagetes erecta

Marigold (Tagetes erecta) is an Asteraceous plant of industrial, ornamental and medicinal importance. This plant is grown as ornamental plant and is adapted to several agro-climates. Its inflorescences have been used as a pigment source for food colouring, mainly of poultry skin and eggs. Tagetes erecta flowers accumulate high levels of carotenoids, especially lutein as the main pigment, and it makes it an attractive model to study the carotenogenic process. There are many reports on the carotenoid effect in the prevention of certain ocular diseases, ischemic heart disease, strokes, photoprotection, immune response, aging and cancer. The aim of this study was to analyze ultra-structural changes occurring during callus development to determine the conditions for the organogenesis process. Marigold calli were induced from leaf explants and they were maintained in MS medium with 2, 4-D and BA (2 mg L ^-1). Leaf explants were grown in MS medium with IAA and BA at different concentrations to evaluate tissue responses, such as production of explants with buds, buds per explant and bud formation capacity. Bud formation was observed using 3.0 mg L^-1 IAA and 0.5 mg L^-1 BA. Subcellular studies on Tagetes erecta calli indicated structures, such as cell walls, nuclei, nuclear membranes, intercellular spaces, chloroplasts, vacuoles and chromoplasts. After tissue culture and subcellular analysis, pBI426 was used to establish bombardment conditions and transient GUS gene expression was achieved with a high pressure gun technique and 70% positive events were recorded. These results will help develop stable callus transformation so that the carotenoid pathway in Tagetes erecta can be modified.