Transformation of Panax notoginseng saponins by steaming and Trichoderma longibrachiatum

Panax notoginseng has been used for medicinal purposes in China for many years. Saponins are believed to be the major bioactive ingredients in P. notoginseng. Two different processes, steaming and biotransformation, were used to study the transformations of saponins in P. notoginseng. During an 8-h steaming process, the ginsenosides Rb1, Rd, Rg1, Re and notoginsenoside R1, decreased to 1.07 mg/g dry weight (DW), 0.91, 0.64, 0 and 0 mg/g DW, respectively. Meanwhile, ginsenoside 20(S)-Rg3, 20(S)-Rh1, F2 and compound K significantly increased to 5.85, 6.10, 0.81 and 6.62 mg/g DW, respectively. On the other hand, one fungus was isolated from the root of P. notoginseng, which could transform ginsenoside Rb1 to ginsenoside Rd specifically. The fungus was identified as Trichoderma longibrachiatum species based on sequence analysis of the rRNA internal transcribed spacers region. The results implied a prospective feasibility for setting up different processing techniques to improve the quality of P. notoginseng and add its value

Strengthening Triterpene Saponins Biosynthesis by Over-Expression of Farnesyl Pyrophosphate Synthase Gene and RNA Interference of Cycloartenol Synthase Gene in Panax notoginseng Cells

To conform to the multiple regulations of triterpene biosynthesis, the gene encoding farnesyl pyrophosphate synthase (FPS) was transformed into Panax notoginseng (P. notoginseng) cells in which RNA interference (RNAi) of the cycloartenol synthase (CAS) gene had been accomplished. Transgenic cell lines showed both higher expression levels of FPS and lower expression levels of CAS compared to the wild-type (WT) cells. In the triterpene and phytosterol analysis, transgenic cell lines provided a higher accumulation of total triterpene saponins, and a lower amount of phytosterols in comparison with the WT cells. Compared with the cells in which RNAi of the CAS gene was achieved, the cells with simultaneously over-expressed FPS and silenced CAS showed higher triterpene contents. These results demonstrate that over-expression of FPS can break the rate-limiting reaction catalyzed by FPS in the triterpene saponins biosynthetic pathway; and inhibition of CAS expression can decrease the synthesis metabolic flux of the phytosterol branch. Thus, more precursors flow in the direction of triterpene synthesis, and ultimately promote the accumulation of P. notoginseng saponins. Meanwhile, silencing and over-expressing key enzyme genes simultaneously is more effective than just manipulating one gene in the regulation of saponin biosynthesis

Interaction of gallic acid with trypsin analyzed by spectroscopy

The interactions between trypsin and gallic acid (GA) were investigated by means of fluorescence spectroscopy, UV-vis absorption spectroscopy, resonance light scattering (RLS) spectroscopy, synchronous fluorescence spectroscopy, and enzymatic inhibition assay. It was found that GA can cause the fluorescence quenching of trypsin during the process of formation of GA-trypsin complex, resulting in inhibition of trypsin activity (IC50 = 3.9 × 10−6 mol/L). The fluorescence spectroscopic data showed that the quenching efficiency can reach about 80%. The binding constants were 1.9371 × 104 L/mol, 1.8192 × 104 L/mol, and 1.7465 × 104 L/mol at three temperatures, respectively. The thermodynamic parameters revealed that hydrogen bonds, van der Waals, hydrophobic, and electrostatic interactions were involved in the binding process of GA to trypsin. Molecular modeling studies illustrated a specific display of binding information and explained most of the experiment phenomena. The microenvironments of tryptophan and tyrosine residue in trypsin were changed by the GA. Results indicated that GA was a strong quencher and inhibitor of trypsin

Molecular Cloning and Characterization of PnbHLH1 Transcription Factor in Panax notoginseng

Panax notoginseng has been extensively used as a traditional Chinese medicine. In the current study, molecular cloning and characterization of PnbHLH1 transcription factor were explored in Panax notoginseng. The full length of the PnbHLH1 gene obtained by splicing was 1430 bp, encoding 321 amino acids. Prokaryotic expression vector pET-28a-PnbHLH1 was constructed and transferred into the BL21 prokaryotic expression strain. An electrophoretic mobility shift assay of PnbHLH1 protein binding to E-box cis-acting elements verified that PnbHLH1 belonged to the bHLH class transcription factor which could interact with the promoter region of the E-box core sequence. The expression levels of key genes involved in the biosynthesis of triterpenoid saponins in PnbHLH1 transgenic cells were higher than those in the wild cells. Similarly, the total saponin contents were increased in the PnbHLH1 transgenic cell lines compared with the wild cell lines. Such results suggest that the PnbHLH1 transcription factor is a positive regulator in the biosynthesis of triterpenoid saponins in Panax notoginseng

Elevated Levels of MYB30 in the Phloem Accelerate Flowering in Arabidopsis through the Regulation of FLOWERING LOCUS T

In Arabidopsis thaliana, the R2R3 MYB-like transcription factor MYB30 is a positive regulator of the pathogen-induced hypersensitive response and of brassinosteroid and abscisic acid signaling. Here, we show that MYB30 expressed under the control of the strong phloem-specific SUC2 promoter accelerates flowering both in long and short days. Early flowering is mediated by elevated expression of flowering locus T (FT), which can be observed in the absence and presence of CONSTANS (CO), the main activator of FT. CO-independent activation by high MYB30 expression results in FT levels that remain below those observed in the wild-type plants, which show an additive CO-dependent activation. In contrast, twin sister of FT (TSF) is repressed in plants expressing high levels of MYB30 in the phloem. In transient assays, MYB30 and CO additively increase the activity of a reporter construct driven by a 1 kb FT promoter. Acceleration of flowering by MYB30 does not require the presence of salicylic acid and is independent of FLC. Taken together, increased levels of MYB30, which was reported to be induced in response to the perception of pathogens, can accelerate flowering and MYB30 may thus be a candidate to mediate cross-talk between gene networks involved in biotic stress perception and flowering time

Additional file 4 of Panax notoginseng transcription factor WRKY15 modulates resistance to Fusarium solani by up-regulating osmotin-like protein expression and inducing JA/SA signaling pathways

Supplementary Material

Genetic and molecular regulation of increased photosynthetic cell number contributes to leaf size heterosis in Arabidopsis

Summary: Heterosis is an important genetic phenomenon that has been observed and widely utilized in agriculture. However, the genetic and molecular bases of heterosis are unclear. Through transcriptome-wide association studies (TWAS) and expression quantitative trait locus (eQTL) analysis to integrate genome, transcriptome, and heterotic phenotype of a half-sibling Arabidopsis hybrid population, we report that the genetic and molecular bases of variations in leaf growth heterosis can be explained by the varied expression levels of growth-regulating genes resulting from distinct sets of heterozygous eQTLs carried by the half-sibling hybrids. In F1 versus parent, the degree of up-regulated gene expression in the cell cycle pathway in the shoot apex and the photosynthesis pathway in true leaf positively correlates with true leaf area heterosis level, and this is affected by the accumulation of superior heterozygous eQTLs. This was further corroborated by the major contribution of increased photosynthetic cell number to leaf area heterosis

Additional file 2 of Panax notoginseng transcription factor WRKY15 modulates resistance to Fusarium solani by up-regulating osmotin-like protein expression and inducing JA/SA signaling pathways

Supplementary Material