Exogenous gibberellic acid shortening after-ripening process and promoting seed germination in a medicinal plant Panax notoginseng.

BACKGROUND: Panax notoginseng (Burk) F.H. Chen is an essential plant in the family of Araliaceae. Its seeds are classified as a type of morphophysiological dormancy (MPD), and are characterized by recalcitrance during the after-ripening process. However, it is not clear about the molecular mechanism on the after-ripening in recalcitrant seeds. RESULTS: In this study, exogenous supply of gibberellic acid (GA3) with different concentrations shortened after-ripening process and promoted the germination of P. notoginseng seeds. Among the identified plant hormone metabolites, exogenous GA3 results in an increased level of endogenous hormone GA3 through permeation. A total of 2971 and 9827 differentially expressed genes (DEGs) were identified in response to 50 mg L-1 GA3 (LG) and 500 mg L-1 GA3 (HG) treatment, respectively, and the plant hormone signal and related metabolic pathways regulated by GA3 was significantly enriched. Weighted gene co-expression network analysis (WGCNA) revealed that GA3 treatment enhances GA biosynthesis and accumulation, while inhibiting the gene expression related to ABA signal transduction. This effect was associated with higher expression of crucial seed embryo development and cell wall loosening genes, Leafy Contyledon1 (LEC1), Late Embryogenesis Abundant (LEA), expansins (EXP) and Pectinesterase (PME). CONCLUSIONS: Exogenous GA3 application promotes germination and shorts the after-ripening process of P. notoginseng seeds by increasing GA3 contents through permeation. Furthermore, the altered ratio of GA and ABA contributes to the development of the embryo, breaks the mechanical constraints of the seed coat and promotes the protrusion of the radicle in recalcitrant P. notoginseng seeds. These findings improve our knowledge of the contribution of GA to regulating the dormancy of MPD seeds during the after-ripening process, and provide new theoretical guidance for the application of recalcitrant seeds in agricultural production and storage.

iTRAQ and RNA-seq analyses provide an insight into mechanisms of recalcitrance in a medicinal plant Panax notoginseng seeds during the after-ripening process.

Panax notoginseng (Burk) F.H. Chen is an important economic and medicinal plant from the family of Araliaceae, and its seed is characterised by the recalcitrance and after-ripening process. However, the molecular mechanism on the dehydration sensitivity is not clear in recalcitrant seeds. In the present study, isobaric tag for relative and absolute quantification (iTRAQ) and RNA-seq were used to analyse the proteomic and transcriptomic changes in seeds of P. notoginseng in days after-ripening (DAR). A total of 454 differentially expressed proteins (DEPs) and 12000 differentially expressed genes (DEGs) were obtained. The activity of enzymes related to antioxidant system were significantly increased, and the late embryogenesis abundant (LEA) protein family and most members of glutathione metabolism enzymes have been downregulated during the after-ripening process. The lack or inadequate accumulation of LEA proteins in the embryo and the low activity of antioxidant defense in glutathione metabolism might be the key factors leading to the dehydration sensitivity in recalcitrant seeds of P. notoginseng. In addition, the increased activity of elycolysis (EMP), citric acid cycle (TCA) and pentose phosphate pathway (PPP) pathways might be one of important signals to complete the after-ripening process. Overall, our study might provide a new insight into the molecular mechanism on dehydration sensitivity of recalcitrant seeds.

Unigene-based RNA-seq provides insights on drought stress responses in Marsdenia tenacissima.

Marsdenia tenacissima is a well-known anti-cancer medicinal plant used in traditional Chinese medicine, which often grows on the karst landform and the water conservation capacity of land is very poorly and drought occurrences frequently. We found M. tenacissima has strong drought resistance because of continuousdrought16 d, the leaves of M. tenacissima were fully curly and dying. But the leaves were fully almost recovering after re-watering 24h. The activity of SOD and POD were almost doubled under drought stress. The content of osmotic regulating substance proline and soluble sugar were three times than control group. But after re-watering, these indexes were declined rapidly. Three cDNA libraries of control, drought stress, and re-watering treatments were constructed. There were 43,129,228, 47,116,844, and 42,815,454 clean reads with Q20 values of 98.06, 98.04, and 97.88respectively.SRA accession number of raw data was PRJNA498187 on NCBI. A total of 8672, 6043, and 6537 differentially expressed genes (DEGs) were identified in control vs drought stress, control vs re-watering, and drought stress vs re-watering, respectively. In addition, 1039, 1016, and 980 transcription factors (TFs) were identified, respectively. Among them, 363, 267, and 299 TFs were identified as DEGs in drought stress, re-watering, and drought stress and re-watering, respectively. These differentially expressed TFs mainly belonged to the bHLH, bZIP, C2H2, ERF, MYB, MYB-related, and NAC families. A comparative analysis found that 1174 genes were up-regulated and 2344 were down-regulated under drought stress and this pattern was the opposite to that found after re-watering. Among the up-regulated genes, 64 genes were homologous to known functional genes that directly protect plants against drought stress. Furthermore, 44 protein kinases and 38 TFs with opposite expression patterns under drought stress and re-watering were identified, which are possibly candidate regulators for drought stress resistance in M. tenacissima. Our study is the first to characterize the M. tenacissima transcriptome in response to drought stress, and will serve as a useful resource for future studies on the functions of candidate protein kinases and TFs involved in M. tenacissima drought stress resistance.

Determination of multiple elements in samples of the medicinal plant Marsdenia tenacissima and estimation of geographic origin via pattern recognition techniques.

Multi-element analysis of the medicinal plant Marsdenia tenacissima was used to develop a reliable method of tracing the geographical source of the samples. The concentrations of 27 elements in 128 samples from 4 provinces in China were analyzed by inductively coupled plasma-atomic emission spectroscopy. Pattern recognition techniques, viz. principal component analysis (PCA), cluster analysis (CA), stepwise linear discriminant analysis (SLDA) and k-nearest neighbor analysis (KNN), were used for this purpose. It was verified that 21 elements in the M. tenacissima samples from different regions showed significant differences (P < 0.05). The PCA explained 87.36 % of the variance with the first seven principal component variables, and a score plot produced from the largest three principal components showed that the source area of most samples could be correctly distinguished. The CA showed that samples were separated into three clusters. The SLDA produced an overall correct classification rate of 87.5 % and a cross-validation rate of 85.2 %. The KNN analysis performed ideally, with an average identification rate of 100 % for the training set and 93.33 % for the test set. These results laid the foundation for the application of multi-element analysis combined with pattern recognition techniques for tracing the geographical origin of samples of medicinal plants.

[Wild resources survey of Marsdenia tenacissima in Honghe, Yunnan].

To ascertain current situation of wild Marsdenia tenacissima resources in Honghe, Yunnan province, the distribution, habitat characteristic and resources reserves of M. tenacissima were surveyed based on interviews and investigation. The results showed that M. tenacissima was found in 7 counties such as Jinping, Mengzi etc, and distributed mainly on the mountainsides from 800 m to 1 200 m. And distribution was affected by many factors, such as light, heat, topography, soil, and vegetation. M. tenacissima grew well in distribution areas. M. tenacissima had averagely a weight of 2.8 kg per plant. Resources reserve of M. tenacissima in Honghe was estimated to 1 300 tons by now but it reduced rapidly in resent years, the wild resources reserve may not meet demand of market. Resources protection and wildlife tending would be conducted to deal with increasing medication requirements.

[Simultaneous determination of chlorogenic acid, scutellarin, 3,5-dicaffeoylquinic acid, 4,5-dicaffeoylquinic acid in different parts of Erigeron breviscapus by high-performance liquid chromatography].

OBJECTIVE: A high-performance liquid chromatographic (HPLC) method was developed for simultaneous determination of chlorogenic acid, scutellarin, 3,5-dicaffeoylquinic acid, 4,5-dicaffeoylquinic acid in different parts of Erigerontis Herba. METHOD: The four constituents were measured on an Agilent Zorbax SB-C18 column (4.6 mm x 450 mm, 5 microm) with a gradient elution of acetonitrile (A) -0.3% phosphoric acid solution (B) (0-10 min, 12%-15% A, 10-32 min, 15% A, 32-33 min, 15%-20% A, 33-50 min, 20%-22% A) at wavelength of 335 nm and 327 nm, and a flow rate of 1.0 mL x min(-1) and the column temperature was 30 degrees C. RESULT: Linearity of each standard was established in the concentration range of 0.050 1-1.002 microg for chlorogenic acid, 0.165 9-3.318 microg for chlorogenic acid, 0.049 7-0.994 microg for 3,5-dicaffeoylquinic acid, 0.048 7-0.974 p.g for 4,5-dicaffeoylquinic acid respectively, with correlation coefficient r > 0.999 6. Average recoveries (n = 6) of 4 compounds were 98.53% with a RSD of 0.94%, 99.68% with a RSD of 0.49%, 98.78% with a RSD of 1.1%, 99.06% with a RSD of 0.81%, respectively. CONCLUSION: The developed method is simple, accurate, and precise, it can be used for the quantitative analysis of Erigeron breviscapus.

[Study on HPLC characteristic fingerprint of Yunnan Dipsacus asper].

OBJECTIVE: To study the HPLC fingerprint of yunnan Dipsacus asper and provide a reliable method for sciencetific evaluation and quality control. METHODS: 16 batches of Dipsacus asper were collected from 11 counties (cities, districts) of 5 prefectures or municipals in Yunnan provinice, which were the mainly distribution region of Dipsacus asper. Samples were analyzed on an Aglient Zorbax SB-18 (4.6 mm x 250 mm, 5 microm) eluted with methanol (A) and water (B) as mobile phase in gradient clution. The flow rate was 1 ml/min, and the column temperature was set at 38 degrees C. The detector wavelength was 220 nm. RESULTS: A HPLC fingerprint method was established, 14 common peaks were selected and the similarity ranged from 0.674 to 0.965, cluster analysis could classified these 16 batches of Dipsacus asper into 2 groups. CONCLUSION: Dipsacus japonicus is found for the first time in yunnan. The fingerprint of Dipsacus asper in different origin have a extremely high similarity;The method is accurate and reliable.