Comprehensive Identification and Profiling of miRNAs Involved in Terpenoid Synthesis of Gleditsia sinensis Lam.

Gleditsia sinensis Lam. is a tree with worldwide distribution and important economic and medicinal values; its pods contain terpenoids including gleditsioside, thiamine, and brassinosteroids. However, thus far, there are few studies on the terpenoid regulation of G. sinensis at the molecular level. microRNA (miRNA) is a class of small RNAs with conserved and crucial roles in the regulation of diverse biological processes during plant growth and development. To identify the miRNAs of G. sinensis and evaluate their involvement in terpenoid synthesis, this investigation quantified the content changes in saponins in pods at three developmental stages: May (pod-setting stage), July (elongation stage), and September (browning stage), and then we performed genome-wide miRNA profiles during the three development stages of the G. sinensis pods. A total of 351 conserved miRNAs belonging to 216 families were identified, among which 36 conserved miRNAs exist specifically in legumes. Through target analysis, 708 unigenes were predicted to be candidate targets of 37 differentially expressed miRNAs. The targets of miR838-3p and miR2093-5p were involved in the derived branches of monoterpenes and gleditsioside, in brassinosteroid biosynthesis (BRB), and in indole alkaloid biosynthesis (IAB). Intriguingly, the targets of miR829-3p.1 were predicted to take part in thiamine biosynthesis, and the targets of miR4414b and miR5037a were involved in the main process of cytokinin synthesis. The corresponding targets participated in BRB, IAB, and terpenoid backbone biosynthesis, which were enriched significantly, suggesting that miR2093-5p, miR4414b, miR5037a, miR829-3p.1, and miR838-3p play indispensable roles in the regulation of triterpenoid saponin and monoterpenoid biosynthesis. To date, this is the first report of miRNA identification in G. sinensis and miRNA expression profiles at different developmental stages of G. sinensis pods, which provides a basis for further uncovering the molecular regulation of terpenoid synthesis in G. sinensis and new insights into the role of miRNAs in legumes.

The Garbage Enzyme with Chinese Hoenylocust Fruits Showed Better Properties and Application than When Using the Garbage Enzyme Alone

Garbage enzyme (GE) is a vinegar or alcohol product derived from fermenting fresh kitchen waste, such as vegetable and fruit residues (peels, cuttings and bits), sugar (brown sugar, jaggery or molasses sugar) and water. Chinese honeylocust fruits (Gleditsia sinensis) have been used in China for at least 2000 years as a detergent. The aim of the study was to investigate the properties and application of Chinese honeylocust garbage enzyme (CHGE), which is produced when equal amounts of Chinese honeylocust fruits and fresh wastes are mixed. The results showed that CHGE had lesser microbial communities and lower surface tension than GE. CHGE also had higher viscosity, foam stability and emulsion stability than GE. Compared with GE, CHGE induced higher enzymatic amylase, cellulase, lipase and protease activities. CHGE had stronger detergency than GE and a 100× dilution of CHGE could significantly remove pesticide residues after a 30 min soaking treatment. The study showed that as a biological detergent, CHGE is safer and more environmentally friendly than GE and has remarkable washing and cleaning power. The preparation method of the detergent is simple: it can be prepared at home using fruit and vegetable waste, which is beneficial to the secondary utilization of waste and the reduction of pollution to the environment and damage to human health.

Effects of Exogenous Calcium on Adaptive Growth, Photosynthesis, Ion Homeostasis and Phenolics of Gleditsia sinensis Lam. Plants under Salt Stress

Salinity is the main environmental factor responsible for limited plant growth in many areas of the world. Gleditsia sinensis Lam. is a shelter forest tree species that does not require high-quality soil and can even grow in mild saline soil. This study mainly explored the tolerance of G. sinensis to salt and the effect of exogenous calcium addition on the growth of G. sinensis in a salinized soil. The concentrations of NaCl were set as 0 mmol/L, 100 mmol/L, and 200 mmol/L. Compared with the control, under the NaCl treatment of 200 mmol/L, it was observed that the leaves of G. sinensis turned yellow, the electrical conductivity significantly increased, and the water content and the chlorophyll content significantly decreased, which is probably unfavorable for growth. Our study showed that the addition of 10 mmol/L exogenous calcium chloride under salt stress had a positive effect on the growth and photosynthetic characteristics of G. sinensis. Moreover, the addition of exogenous calcium attenuated the cytotoxicity caused by Na+ under salt stress and promoted the equilibrium of ion homeostasis. More importantly, the addition of exogenous calcium ions was beneficial for the survival of G. sinensis plants on salinized land and the increase of effective active ingredient content including phenolic compounds, which is of direct significance for improving environmental problems such as desertification of saline-alkali land. In conclusion, we investigated the effect of salt treatment on G. sinensis, as well as the positive effects of exogenous calcium on the survival and growth of G. sinensis in salt environment, which provided a scientific basis for the targeted cultivation of G. sinensis in salinized land and the effective utilization of salinized and alkaline land.

A High-Content Screen for the Identification of Plant Extracts with Insulin Secretion-Modulating Activity

Bioactive plant compounds and extracts are of special interest for the development of pharmaceuticals. Here, we describe the screening of more than 1100 aqueous plant extracts and synthetic reference compounds for their ability to stimulate or inhibit insulin secretion. To quantify insulin secretion in living MIN6 β cells, an insulin–Gaussia luciferase (Ins-GLuc) biosensor was used. Positive hits included extracts from Quillaja saponaria, Anagallis arvensis, Sapindus mukorossi, Gleditsia sinensis and Albizia julibrissin, which were identified as insulin secretion stimulators, whereas extracts of Acacia catechu, Myrtus communis, Actaea spicata L., Vaccinium vitis-idaea and Calendula officinalis were found to exhibit insulin secretion inhibitory properties. Gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS) were used to characterize several bioactive compounds in the selected plant extracts, and these bioactives were retested for their insulin-modulating properties. Overall, we identified several plant extracts and some of their bioactive compounds that may be used to manipulate pancreatic insulin secretion.

Insights into molecular structure, genome evolution and phylogenetic implication through mitochondrial genome sequence of Gleditsia sinensis

Gleditsia sinensis is an endemic species widely distributed in China with high economic and medicinal value. To explore the genomic evolution and phylogenetic relationships of G. sinensis, the complete mitochondrial (mt) genome of G. sinensis was sequenced and assembled, which was firstly reported in Gleditsia. The mt genome was circular and 594,121 bp in length, including 37 protein-coding genes (PCGs), 19 transfer RNA (tRNA) genes and 3 ribosomal RNA (rRNA) genes. The overall base composition of the G. sinensis mt genome was 27.4% for A, 27.4% for T, 22.6% for G, 22.7% for C. The comparative analysis of PCGs in Fabaceae species showed that most of the ribosomal protein genes and succinate dehydrogenase genes were lost. In addition, we found that the rps4 gene was only lost in G. sinensis, whereas it was retained in other Fabaceae species. The phylogenetic analysis based on shared PCGs of 24 species (22 Fabaceae and 2 Solanaceae) showed that G. sinensis is evolutionarily closer to Senna species. In general, this research will provide valuable information for the evolution of G. sinensis and provide insight into the phylogenetic relationships within the family Fabaceae.

Identification of the initial water-site and movement in Gleditsia sinensis seeds and its relation to seed coat structure

Background Water uptake is essential for seed germination. However, Gleditsia sinensis seeds have a water-impermeable seed coat, which is beneficial for its adaption to the environment, but prohibits its germination without treatment. This feature may be associated with the structure of the seed coat. Thus, the aim of this research was to identify and describe the initial water uptake site and water movement and to determine the relationship between seed coat structure and water absorption. Results A water temperature of 80 °C was optimal to break the hardseededness of G. sinensis seeds. Scanning electron microscopy (SEM) images revealed that the seed coat consisted of a palisade layer and light line that can hinder water entry into the seed. Also, a structure of vascular bundles existed in the hilar region. Hot water treatment caused the tightly closed micropyle to open and the cavity beneath it expanded; the layer of palisade cells in the lens was raised. The embryo dye-tracking tests showed that the radicle tip was the initial region to be stained red. After staining for 24 h, the red-stained area on the vascular bundle side of cotyledon was more extensive than that on the other side. Further studies by MRI maps indicated that the micropyle was the initial site for water imbibition. Some water then migrated along the space between the seed coat and the endosperm to the chalazal; simultaneously, the rest of the water reached the embryonic axis and spread into cotyledons. The maps of 20–24 h showed that water was unevenly distributed within the cotyledons in a way that the edge parts were more hydrated than the center. Blocking tests showed that the hilar region was the initial and an important region during seed imbibition. The medial region and chalazal portion were capable of imbibing water when the hilar region was blocked, but water absorption was later and slower than that through the hilar region. Conclusion MRI technology provides a promising and non-invasive technique to identify the water gap and the path of water movement in the seed. Combined with the results of SEM, the relation between seed coat and its imbibition can be inferred.