Metabolome and transcriptome reveal the biosynthesis of flavonoids and amino acids in Isatis indigotica fruit during development

Isatis indigotica Fort. is a famous medicinal plant that is also used as a natural dye and functional vegetable. The characteristics of the I. indigotica fruit during development are largely unknown, information that is essential for the exploitation and seedlings cultivation of I. indigotica. In this study, the biochemical, metabolite characteristics and gene expression profiling of I. indigotica at four developmental stages were investigated. A total of 428 metabolites were detected and categorized into 17 categories. High contents of anthocyanins, especially cyanidin 3-glucoside, might contribute to the purple colouration of I. indigotica fruits. Moreover, dozens of flavonoid components, including taxifolin, quercetin, astragalin and isovitexin 2″-O-beta-D-glucoside, and several other active components were also up-regulated in mature fruits. The abundance of antioxidants might endow a significantly stronger antioxidant activity of mature I. indigotica fruits compared to many other reported species. Enrichment analyses revealed that flavonoid and anthocyanin biosynthesis genes were mostly enriched in up-regulated gene sets during fruit development. The up-regulated structural genes, including IiCHS, IiCHI, IiF3H, IiDFR, IiANS, IiFLS, IiUGT, and transcription factors such as IiMYBs, IibHLHs and IiNACs were identified as candidate regulators of flavonoid and anthocyanin biosynthetic pathway. Furthermore, biosynthesis of amino acids was enriched in all pairwise comparisons of metabolites in fruits at four developmental stages. The differential accumulation of amino acids might result from the differentially expressed genes involved in amino acid biosynthesis. Taken together, these findings provide a comprehensive understanding of metabolite profiling and gene expression patterns in I. indigotica fruit during maturity, which is useful for pharmaceutical extractions and seedling cultivation of I. indigotica.