Studies of genes involved in the metabolic steps of indigo production in Isatis tinctoria

Plant stain utilisation is well documented from ancient times: British Picti represent a historical known case of a people using skin decoration with indigo extracted from the woad plant Isatis tinctoria L. The present work aims at the genetic improvement of plants producing the indigo stain and to the re-introduction of plants yielding high indigo. In this direction, we are interested to individuate the genes involved in the metabolic pathway of indigo biosynthesis in Isatis tinctoria L., for their use in assisted selection programs. We have hypothesised candidates genes involved in the metabolic pathways and we have designed degenerated primers for the amplification and isolation of these genes. At the moment, we are studying the alfa-tryptophanase gene involved in the the first step of indican biosynthesis, the indigo precursor normally stored in vacuoli, and the beta-D-glucosidase gene involved in indican hydrolysis leading to the release of indoxyl groups and indigo synthesis. In this context, we have isolated plants with high indigo production, we have amplified the beta-D-Glucosidase and the alfa-Tryptophanase genes. Moreover, the expression of these genes during different developmental stages is under study through analysis of mRNA biosynthesis. This work is carried out within the EC Spindigo Project.