Single cell subtractive transcriptomics for identification of cell-specifically expressed candidate genes of pyrrolizidine alkaloid biosynthesis

摘要: Progress has recently been made in the elucidation of pathways secondary metabolism. However, because its diversity, genetic information concerning biosynthetic details is still missing for many natural products. This also case biosynthesis pyrrolizidine alkaloids. To close this gap, we tested strategies using tissues that express pathway comparison to which not expressed. As metabolism are known be induced by jasmonates, alkaloid-producing species Heliotropium indicum, Symphytum officinale, and Cynoglossum officinale Boraginales order were treated with methyl jasmonate. An effect on alkaloid levels transcript homospermidine synthase, first specific enzyme biosynthesis, was detectable. Therefore, a method developed making use often observed cell-specific production compounds. H. indicum produces alkaloids exclusively shoot. Homospermidine synthase expressed only cells lower leaf epidermis stem. Suggesting whole might localized these cells, have isolated single upper laser-capture microdissection. The resulting cDNA preparations used subtractive transcriptomic approach. Quantitative real-time polymerase chain reaction shown library significantly enriched homospermidine-synthase-coding transcripts providing valuable source identification further genes involved biosynthesis.