Polygonati Rhizoma demonstrates significant potential for addressing both chronic and hidden hunger. The supply of high-quality seedlings is a primary factor influencing the development of the Polygonati Rhizoma industry. Warm water soaking is often used in agriculture to promote the rapid germination of seeds, while its application and molecular mechanism in Polygonati Rhizoma have not been reported. To rapidly obtain high-quality seedlings, this study treated Polygonatum kingianum var. grandifolium seeds with sand storage at low temperatures, warm water soaking, and cultivation temperature gradients. The results showed that the culture at 25 ℃ or sand storage at 4 ℃ for 2 months rapidly broke the seed dormancy of P. kingianum var. grandifolium, while the culture at 20 ℃ or sand storage at 4 ℃ for 1 month failed to break the seed dormancy. Soaking seeds in 60 ℃ warm water further increased the germination rate, germination potential, and germination index. Specifically, the seeds soaked at 60 ℃ and cultured at 25 ℃ without sand storage treatment(Aa25) achieved a germination rate of 78. 67%±1. 53% on day 42 and 83. 40%±4. 63% on day 77. The seeds pretreated with sand storage at 4 ℃ for 2 months, soaked in 60 ℃ water, and then cultured at 25 ℃ achieved a germination rate comparable to that of Aa25 on day 77. Transcriptomic analysis indicated that warm water soaking might promote germination by triggering reactive oxygen species( ROS), inducing the expression of heat shock factors( HSFs) and heat shock proteins( HSPs), which accelerated DNA replication, transcript maturation, translation, and processing, thereby facilitating the accumulation and turnover of genetic materials. According to the results of indoor controlled experiments and field practices, maintaining a germination and seedling cultivation environment at approximately 25 ℃ was crucial for the one-year seedling cultivation of P. kingianum var. grandifolium.
Germination ; Seedlings/genetics* ; Water/metabolism* ; Seeds/metabolism* ; Polygonatum/genetics* ; Temperature ; Plant Proteins/genetics* ; Plant Dormancy

This study aims to investigate the protective effects and mechanisms of polysaccharide extract PCP1 from Polygonatum cyrtonema in ameliorating cerebral ischemia-reperfusion(I/R) injury in rats through modulation of the Toll-like receptor 4(TLR4)/NOD-like receptor protein 3(NLRP3) signaling pathway. In vivo, SD rats were randomly divided into the sham group, model group, PCP1 group, nimodipine(NMDP) group, and TLR4 signaling inhibitor(TAK-242) group. A middle cerebral artery occlusion/reperfusion(MCAO/R) model was established, and neurological deficit scores and infarct size were evaluated 24 hours after reperfusion. Hematoxylin-eosin(HE) and Nissl staining were used to observe pathological changes in ischemic brain tissue. Transmission electron microscopy(TEM) assessed ultrastructural damage in cortical neurons. Enzyme-linked immunosorbent assay(ELISA) was used to measure the levels of interleukin-1β(IL-1β), interleukin-6(IL-6), interleukin-18(IL-18), tumor necrosis factor-α(TNF-α), interleukin-10(IL-10), and nitric oxide(NO) in serum. Immunofluorescence was used to analyze the expression of TLR4 and NLRP3 proteins. In vitro, a BV2 microglial cell oxygen-glucose deprivation/reperfusion(OGD/R) model was established, and cells were divided into the control, OGD/R, PCP1, TAK-242, and PCP1 + TLR4 activator lipopolysaccharide(LPS) groups. The CCK-8 assay evaluated BV2 cell viability, and ELISA determined NO release. Western blot was used to analyze the expression of TLR4, NLRP3, and downstream pathway-related proteins. The results indicated that, compared with the model group, PCP1 significantly reduced neurological deficit scores, infarct size, ischemic tissue pathology, cortical cell damage, and the levels of inflammatory factors IL-1β, IL-6, IL-18, TNF-α, and NO(P<0.01). It also elevated IL-10 levels(P<0.01) and decreased the expression of TLR4 and NLRP3 proteins(P<0.05, P<0.01). Moreover, in vitro results showed that, compared with the OGD/R group, PCP1 significantly improved BV2 cell viability(P<0.05, P<0.01), reduced cell NO levels induced by OGD/R(P<0.01), and inhibited the expression of TLR4-related inflammatory pathway proteins, including TLR4, myeloid differentiation factor 88(MyD88), tumor necrosis factor receptor-associated factor 6(TRAF6), phosphorylated nuclear factor-kappaB dimer RelA(p-p65)/nuclear factor-kappaB dimer RelA(p65), NLRP3, cleaved-caspase-1, apoptosis-associated speck-like protein(ASC), GSDMD-N, IL-1β, and IL-18(P<0.05, P<0.01). The protective effects of PCP1 were reversed by LPS stimulation. In conclusion, PCP1 ameliorates cerebral I/R injury by modulating the TLR4/NLRP3 signaling pathway, exerting anti-inflammatory and anti-pyroptotic effects.
Animals ; Toll-Like Receptor 4/genetics* ; NLR Family, Pyrin Domain-Containing 3 Protein/genetics* ; Rats, Sprague-Dawley ; Rats ; Reperfusion Injury/genetics* ; Male ; Signal Transduction/drug effects* ; Polysaccharides/isolation & purification* ; Polygonatum/chemistry* ; Brain Ischemia/genetics* ; Drugs, Chinese Herbal/administration & dosage* ; Mice ; Humans

Rhizome rot disease is one of the main disease of planted Polygonatum kingianum. In this study, six strains of pathogenic fungus was isolated from P. kingianum samples with rhizome rot disease collected from six counties in Yunnan province. Its pathogenicity was confirmed by inoculation to healthy P. kingianum rhizome according to Koch's postulates. The colonies of the isolated fungi on potato dextrose agar(PDA) were orange with abundant crescentic conidia which were eseptate with a mean size of 19. 3-24. 9 μm×5. 2-5. 9 μm and a L/W ratio of 3. 4-4. 5. There was an oil ball in the center of the conidium. It's easy to see setae on PDA colony.The phylogenetic tree based on ITS, GAPDH, CHS-1, HIS3, ACT, and TUB2 sequences by maximum likelihood(ML) method indicated that the pathogenic fungus for P. kingianum rhizome rot disease was clustered into the clade of Colletotrichum spaethianum species complex, and was close to C. spaethianum. However, there were some differences in morphological and genetic characteristics between the pathogenic fungus and C. spaethianum. Therefore, the pathogenic fungus for rhizome rot disease of P. kingianum was identified as a new Colletotrichum species named C. kingianum. The disease spreads primarily due to the plantation of infected seedlings of P. kingianum. It is necessary to choose healthy seedlings and take rigorous disinfection measures for the disease prevention.
China ; Colletotrichum/genetics* ; Phylogeny ; Polygonatum ; Rhizome

The genomic DNA were extracted from the leaves of Polygonatum roseum (Liliaceae) in Xinjiang and the primers were designed according to conservative sequences of Polygonatum lectins gene. The complete ORF of Polygonatum roseum agglutinin (PRA) gene was amplified as a fragment of 550 bp, which was identical with predicted size. Like most of the plant lectin genes, there was no intron in the PRA gene. The ORF of the gene encoded 159 amino acid residues, in which included a signal sequence of 28 amino acid residues at its N-terminus. The cDNA sequence had 92% identities compared with the published sequence. The amino acid sequence and SWISS-MODEL analysis indicated that the three-dimensional structure of PRA strongly resembled with that of monocot mannose-binding lectins, which comprised with three antiparallel four-stranded beta-sheets arranged as a 12-stranded beta-barrel. The recombinant pGEX4T-1-PRA and pMAL-p2x-PRA prokaryotic expression vectors were constructed to produce GST-PRA and MBP-PRA fusion proteins in E. coli, respectively. SDS-PAGE of the fusion protein demonstrated that the PRA lectin protein migrated at a size of 14 kD. The immunization was performed by intra-muscular injection of pcDNA3-PRA, and the antiserum was detected by ELISA. Western blotting analysis showed the antiserum specifically bound the lectin protein. The establishment of such an expression system might provide materials for further investigation of the properties and functions of PRA proteins. It also laid the basis for plant genetic engineering on its defensive functions to pests and diseases.
Amino Acid Sequence ; Animals ; Base Sequence ; China ; Cloning, Molecular ; Escherichia coli ; genetics ; metabolism ; Female ; Mannose-Binding Lectin ; biosynthesis ; genetics ; Mice ; Molecular Sequence Data ; Polygonatum ; chemistry ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; pharmacology ; Sequence Analysis