Mi Jin Jeon

Caulophyllum robustum, commonly named Asian blue cohosh, is a perennial herb in the family Berberidaceae. It has traditionally been used for folk medicine in China. We isolated berberine from the leaves, stem, roots, and fruits of C. robustum, and this is the first report on berberine in this species. Transcriptome analysis was conducted for the characterization of berberine biosynthesis genes in C. robustum, in which, all the genes for berberine biosynthesis were identified. From 40,094 transcripts, using gene ontology (GO) analysis, 26,750 transcripts were assigned their functions in the categories of biological process, molecular function, and cellular component. In the analysis of genes expressed in different tissues, the numbers of genes in the categories of intrinsic component of membrane and transferase activity were up-regulated in leaves versus stem. The berberine synthesis genes in C. robustum were characterized by phylogenetic analysis with corresponding genes from other berberine-producing species. The co-existence of genes from different plant families in the deepest branch subclade implies that the differentiation of berberine synthesis genes occurred early in the evolution of berberine-producing plants. Furthermore, the copy number increment of the berberine synthesis genes was detected at the species level.

The annual herb Euphorbia maculata L. produces anti-inflammatory and biologically active substances such as triterpenoids, tannins, and polyphenols, and it is used in traditional Chinese medicine. Of these bioactive compounds, terpenoids, also called isoprenoids, are major secondary metabolites in E. maculata. Full-length cDNA sequencing was carried out to characterize the transcripts of terpenoid biosynthesis reference genes and determine the copy numbers of their isoforms using PacBio SMRT sequencing technology. The Illumina short-read sequencing platform was also employed to identify differentially expressed genes (DEGs) in the secondary metabolite pathways from leaves, roots, and stems. PacBio generated 62 million polymerase reads, resulting in 81,433 high-quality reads. From these high-quality reads, we reconstructed a genome of 20,722 genes, in which 20,246 genes (97.8%) did not have paralogs. About 33% of the identified genes had two or more isoforms. DEG analysis revealed that the expression level differed among gene paralogs in the leaf, stem, and root. Whole sets of paralogs and isoforms were identified in the mevalonic acid (MVA), methylerythritol phosphate (MEP), and terpenoid biosynthesis pathways in the E. maculata L. The nucleotide information will be useful for identifying orthologous genes in other terpenoid-producing medicinal plants.

Berberis koreana is a medicinal plant containing berberine, which is a bioactive compound of the benzylisoquinoline alkaloid (BIA) class. BIA is widely used in the food and drug industry for its health benefits. To investigate the berberine biosynthesis pathway, gene expression analysis was performed in leaves, flowers, and fruits at different stages of growth. This was followed by full-length cDNA sequencing analysis using the PacBio sequencer platform to determine the number of isoforms of those expressed genes. We identified 23,246 full-length unigenes, among which 8479 had more than one isoform. The number of isoforms ranged between two to thirty-one among all genes. Complete isoform analysis was carried out on the unigenes encoding BIA synthesis. Thirteen of the sixteen genes encoding enzymes for berberine synthesis were present in more than one copy. This demonstrates that gene duplication and translation into isoforms may contribute to the functional specificity of the duplicated genes and isoforms in plant alkaloid synthesis. Our study also demonstrated the streamlining of berberine biosynthesis via the absence of genes for enzymes of other BIAs, but the presence of all the genes for berberine biosynthesize in B. koreana. In addition to genes encoding enzymes for the berberine biosynthesis pathway, the genes encoding enzymes for other BIAs were not present in our dataset except for those encoding corytuberine synthase (CTS) and berbamunine synthase (BS). Therefore, this explains how B. koreana produces berberine by blocking the pathways leading to other BIAs, effectively only allowing the pathway to lead to berberine synthesis.