Abstract
Baphicacanthus cusia (Nees) Bremek is an herb widely used for the clinical treatment of colds, fever, and influenza in Traditional Chinese Medicine. The roots, stems and leaves can be used as natural medicine, in which indigo and indirubin are two main active ingredients. In this study, quantification of indigo, indirubin, indican and adenosine among various tissues of B. cusia was conducted using HPLC-DAD. Leaves have significantly higher contents than stems and roots (380.66, 315.15, 20,978.26, 4323.15 μg/g in leaves, 306.36, 71.71, 3,056.78, 139.45 μg/g in stems, and 9.31, 7.82, 170.45, 197.48 μg/g in roots, respectively). De novo transcriptome sequencing of B. cusia was performed for the first time. The sequencing yielded 137,216,248, 122,837,394 and 140,240,688 clean reads from leaves, stems and roots respectively, which were assembled into 51,381 unique sequences. A total of 33,317 unigenes could be annotated using the databases of Nr, Swiss-Prot, KEGG and KOG. These analyses provided a detailed view of the enzymes involved in indican backbone biosynthesis, such as cytochrome P450, UDP-glycosyltransferase, glucosidase and tryptophan synthase. Analysis results showed that tryptophan synthase was the candidate gene involved in the tissue-specific biosynthesis of indican. We also detected sixteen types of simple sequence repeats in RNA-Seq data for use in future molecular mark assisted breeding studies. The results will be helpful in further analysis of B. cusia functional genomics, especially in increasing biosynthesis of indican through biotechnological approaches and metabolic regulation.
Highlights
After total RNA was extracted, mRNA isolated from leaves, stems and roots of B. cusia was enriched by oligo beads in order to acquire a general set of B. cusia transcripts
Our work provides novel information with respect to the mechanism of different tissues of B. cusia used as natural medicine, since this may be related to the distribution of indican and the metabolites indigo and indirubin
A total of 63 unigenes related to nitrogen metabolism pathways and 58 unigenes involved in the biosynthesis pathway of indole and its derivatives were found
Summary
Baphicacanthus cusia (Nees) Bremek (B. cusia) named Strobilanthes cusia is a perennial medicinal herb distributed broadly in Fujian, Zhejiang, Guangdong, Hainan, Guangxi, Hong. To better understand the biosynthesis of indigo, gene expression possibly involved in indigo biosynthesis have been analysed by RNA-Seq in Indigofera tinctoria and Polygonum tinctorium [21], species closely related to B. cusia. These organisms belong to different genera and species classifications and differ in morphological traits. To understand the biosynthesis and metabolism pathways of indican in root, stem and leaf tissues of B. cusia, we generated multi-tissue transcriptomic data of B. cusia using RNA-Seq and annotated the multi-tissue transcriptome using publicly available databases and tools; we ascertained genes that may be related to indican biosynthesis and metabolism pathways. The functional characterization of the B. cusia genes mentioned above should be further investigated
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