ABSTRACT
To screen the genes regulating the biosynthesis of phenolic acid derivatives from the genome of Bletilla striata, we designed a suspension culture system to sample the cells for the following experiments. The contents of four phenolic acid derivatives were determined by high-performance liquid chromatography, and several full-length transcriptome sequencings of RNA samples at 10 time points were performed for bioinformatics analysis. The correlation analysis was used to identify and verify the key DEGs involved in the biosynthesis of the four phenolic acid derivatives. The results showed that the contents of p-hydroxybenzylalcohol (HBA), Dactylorhin A, Militarine, and Coelonin peaked at 33 days postinoculation (Dpi), 18 Dpi, 39 Dpi, and 39 Dpi of the culture system, respectively. Based on transcriptome data, 80 DEGs involved in the biosynthesis of phenolic acid derivatives were obtained. The KEGG pathway enrichment analysis classified them mostly into five metabolic pathways: phenylpropane biosynthesis, starch and sucrose metabolic, cyanoamino acid metabolism, gluconeogenesis and glycolysis, and phenylalanine metabolism. qPCR analysis revealed that the relative gene expression levels were consistent with the overall trend of transcriptome sequencing results. Among them, 14, 18, 23, and 41 unigenes were found to be involved in the synthesis of HBA, Dactylorhin A, Coelonin, and Militarine, respectively. These unigenes laid a solid foundation for elucidating the biosynthesis mechanism of phenolic acid derivatives in suspension cells of B. striata.
ABSTRACT
The cultivation medium of Dendrobium nobile has an effect on the contents of its main medicinal components, but the specific mechanism is still unclear. In this study, the callus, seedlings, rhizomes, and leaves of D. nobile were sequenced for the PacBio SMRT. The 2-year-old stems were selected for the Illumina sequencing and metabolome sequencing to analyze the genetic mechanism of metabolic differences under different epiphytic patterns. As a result, a total of 387 differential genes were obtained, corresponding to 66 differential metabolites. Different epiphytic patterns can induce a series of metabolic changes at the metabolome and transcriptome levels of D. nobile, including flavonoid metabolism, purine metabolism, terpenoid backbone biosynthesis, amino acid metabolism, and alpha-linolenic acid metabolic, and related regulatory genes include ALDH2B7, ADC, EPSPS-1, SHKA, DHAPS-1, GES, ACS1, SAHH, ACS2, CHLP, LOX2, LOX2.3, and CYP74B2. The results showed that the genetic mechanism of D. nobile under various epiphytic patterns was different. In theory, the content of metabolites under the epiphytic patterns of Danxia stone is higher, which is more suitable for field cultivation.
ABSTRACT
BACKGROUND: Bletilla striata has been widely used in the pharmacology industry. To effectively produce the secondary metabolites through suspension cultured cells of B. striata, it is important to exploring the full-length transcriptome data and the genes related to cell growth and chemical producing of all culture stages. We applied a combination of Real-Time Sequencing of Single Molecule (SMRT) and second-generation sequencing (SGS) to generate the complete and full-length transcriptome of B. striata suspension cultured cells. METHODS: The B. striata transcriptome was formed in de novo way by using PacBio isoform sequencing (Iso-Seq) on a pooled RNA sample derived from 23 samples of 10 culture stages, to explore the potential for capturing full-length transcript isoforms. All unigenes were obtained after splicing, assembling, and clustering, and corrected by the SGS results. The obtained unigenes were compared with the databases, and the functions were annotated and classified. RESULTS AND CONCLUSIONS: A total of 100,276 high-quality full-length transcripts were obtained, with an average length of 2530 bp and an N50 of 3302 bp. About 52% of total sequences were annotated against the Gene Ontology, 53,316 unigenes were hit by KOG annotations and divided into 26 functional categories, 80,020 unigenes were mapped by KEGG annotations and clustered into 363 pathways. Furthermore, 15,133 long-chain non-coding RNAs (lncRNAs) were detected. And 68,996 coding sequences were identified based on SSR analysis, among which 31 pairs of primers selected at random were amplified and obtained stable bands. In conclusion, our results provide new full-length transcriptome data and genetic resources for identifying growth and metabolism-related genes, which provide a solid foundation for further research on its growth regulation mechanisms and genetic engineering breeding mechanisms of B. striata.
ABSTRACT
Auxin/Indole-3-Acetic Acid (Aux/IAA) genes are involved in auxin signaling pathway and play an important role in plant growth and development. However, many studies focus on Aux/IAA gene families and much less known in Bletilla striata. In this study, a total of 27 Aux/IAA genes (BsIAA1-27) were cloned from the transcriptome of Bletilla striata. Based on a phylogenetic analysis of the Aux/IAA protein sequences from B. striata, Arabidopsis thaliana and Dendrobium officinale, the Aux/IAA genes of B. striata (BsIAAs) were categorized into 2 subfamilies and 9 groups. While BsIAAs were more closer to those of D. officinale compared to A. thaliana. EST-SSR marker mining test showed that 4 markers could be stably amplified with obvious polymorphisms among 4 landraces. Our results suggested that BsIAAs were involved in the process of tuber development and provided insights into functional roles of Aux/IAA genes in B. striata and other plants.
