ABSTRACT
Rhamnose synthase (RHM) is a key enzyme in the biosynthesis of uridine diphosphate rhamnose (UDP-Rha), reversibly converting uridine diphosphate-glucose (UDP-Glc) into UDP-Rha in the presence of NADH or NADPH. In this research, yeast extract (YE) was used to stimulate Sorbus aucuparia suspension cells. Based on a previous study of the transcriptome database of S. aucuparia suspension cells, two RHMs were cloned from S. aucuparia and named SaRHM1 (GenBank No.: MK213340) and SaRHM2 (GenBank No.: MK213341). The SaRHM1 gene contained a 2 007 bp open reading frame (ORF) encoding a polypeptide of 668 amino acids with a molecular weight of 75.25 kD, and a theoretical isoelectric point (pI) of 7.24. The SaRHM2 gene contained a 2 040 bp ORF encoding a polypeptide of 679 amino acids with a molecular weight of 76.26 kD and pI of 6.41. Bioinformatic analysis indicated that SaRHM1 and SaRHM2 contained two special sequences of GxxGxxG/A and YxxxK. Multiple sequence alignments and phylogenetic trees show that SaRHM1 and SaRHM2 have high sequence similarity with other plant species of RHMs. The results of enzyme activity assays in vitro revealed that both recombinant SaRHM1 and SaRHM2 are able to convert UDP-Glc into UDP-Rha. SaRHMs displayed maximum activity at 40 ℃ and a pH of 8 and 9, respectively. The Km values of SaRHM1 and SaRHM2 for UDP-Glc were 212.4 ± 56.70 and 361.0 ± 63.74 μmol·L-1, respectively, with Vmax values of 235.5 ± 18.98 and 516.5 ± 22.30 nmol·min-1·μg-1, respectively. This study reports the cloning and sequencing of RHMs from S. aucuparia and verifies their function, which likely provide rhamnose donors for the subsequent biosynthesis of rhamnosides.
ABSTRACT
Plants have a memory function for the environmental stress they have suffered. When they are subjected to repeated environmental stress, they can quickly and better activate the response and adaptation mechanism to environmental stress, thus realizing long-term stable reproduction. However, most of the relevant studies are applied to crops and Arabidopsis thaliana rather than medicinal plants about the improvement of plant growth status and the effect on phytoalexin biosynthesis. In this study, yeast extract(YE) was used as an elicitor to simulate biotic stress, and the changes in biomass and the content of some secondary metabolites were measured by giving repeated stresses to Sorbus aucuparia suspension cell(SASC). The results showed that the accumulation levels of biomass and some secondary metabolites in SASC subjected to repeated stress are significantly increased at some time points compared with single stress. A phenomenon that SASC can memorize biotic stress is confirmed in this study and influences phytoalexin accumulation in SASC. Furthermore, the work laid the groundwork for research into the transgenerational stress memory mechanism of medicinal plant.
Subject(s)
Cells, Cultured , Secondary Metabolism , Sorbus , Stress, PhysiologicalABSTRACT
Objective: To obtain the glycosyltransferase gene involved in modification reaction of phytoalexin from Sorbus pohuashanensis suspension cell,and conduct sequence analysis and prokaryotic expression analysis. Method: Based on the transcriptome data,specific primers were designed to obtain 2 cDNA sequences of SaUGTs genes,construct prokaryotic expression vector HIS-MBP-pET28a-SaUGTs and induce the expression of recombinant SaUGTs protein. Result: SaUGT1 and SaUGT2 sequences were cloned and obtained from glycosyltransferases,then bioinformatic analysis of the sequence and prokaryotic expression analysis were conducted. SaUGT1 gene contained 1 458 bp open reading frame (ORF),encoding a polypeptide of 485 amino acids,with a relative molecular weight of 54.27 kDa and theoretical isoelectric point (pI) of 5.50.SaUGT2 gene contained 1 431 bp ORF,encoding a polypeptide of 476 amino acids,with a relative molecular weight of 53.49 kDa and theoretical pI of 5.63. Bioinformatics analysis indicated that SaUGT1 and SaUGT2 protein had no signal peptide,and the conserved domains of glycosyltransferase family were detected. Phylogenetic results showed that SaUGT1 and SaUGT2 proteins had the closest relationship with the UGT85 family of A. thaliana. Differential expression analysis revealed that the relative expression levels of SaUGT1 and SaUGT2 were increased significantly after being induced by yeast extract (YE), with the highest expression level found at 24 h and 12 h. The recombinant SaUGT1 and SaUGT2 proteins were successfully expressed in Escherichia coli DE3 cells and finally,the recombinant SaUGT1 and SaUGT2 proteins were purified through Ni2+ affinity chromatography. Conclusion: The glycosyltransferase gene was cloned from the S. aucuparia for the first time,and the prokaryotic expression vector was successfully constructed,laying foundation for further study of the function of this gene.
ABSTRACT
The chemical constituents of plants from the genus Sorbus are mainly flavonoids,biphenyls,terpenoids and organic acids.Pharmacological researches proved that the chemical components isolated from Sorbus plants has antitussive,relieving asthma,antibacterial and antioxidant effect.Moreover,the Sorbus plants were used as landscape trees.There is a bright developing prospect about Sorbus plants.In this review,we give an overview of the significant progress and the latest findings made in Sorbus plants,such as the stress response,chemical compositions,pharmacological activities and the propagation technology.The results including as following:①more evidences are needed for species identification of Sorbus,②the researches about Sorbus plants is focused on only a few species(S. alnifolia,S. amabilis,S. pohuashanensis,S. tianschanica,S. aucuparin),③the existing researches centre upon chemical composition analysis and propagation and cultivation;the formation and mechanism of active components are rarely reported.Thus,the research of Sorbus plants is still in the early period,and this review paper will provide theoretical basis and reference for further research of Sorbus plants.
ABSTRACT
Objective: To explore the functions of pathogenesis related protein 1-like (PR1-like) in suspension cell of Sorbus aucuparia (SaPR1-like) under biotic stress. Method: The full sequence of SaPR1-like gene was cloned and analyzed by multiple bioinformatic tools. The expression of SaPR1-like in the suspension cell of S. aucuparia in response to harpin protein stress was analyzed by real-time fluorescence quantitative polymerase chain reaction (PCR). Result: A SaPR1-like gene was identified and cloned, which contained a complete open reading frame and encoded 161 amino acids. The coding protein was hydrophilic and stable, had transmembrane structure and signal peptide, and belonged to secretory protein. Amino acid sequence alignment data indicated that the C-terminal of SaPR1-like contained a highly conserved domain[cysteine-rich secretory protein, antigen 5, and pathogenesis-related 1 peptides (CAPE)], which could induce defense genes to produce immune responses against biotic stresses. SaPR1-like gene expression was significantly increased after harpin protein induced suspension cell of S. aucuparia for 24 h. Conclusion: A SaPR1-like gene derived from the PR1 family is found to induce significant responses to biotic elicitors in S. aucuparia. This study highlights a role for PR1 in immune signaling and suggests the potential application of PR1 in efforts to defeat biotic stress in plants.
ABSTRACT
Studies on the characteristic chemical markers of sulfur fumigation have become an effective auxiliary way for quality control of traditional Chinese medicine. This study established a quantitative analysis method for the determination of -hydroxybenzyl hydrogen sulfite (-HS) in Gastrodiae Rhizoma (GR) based on UPLC-MS/MS. Then, 37 batches of GR were screened, and the results showed that 27 batches of them were sulfur-fumigated. Especially, the GR samples in Yunnan producing areas were sulfur-fumigated more seriously. Based on the stability of -HS after different storage time and heat treatment methods, it was found that the content of -HS was stable and reliable in the storage period of 8 months and under normal heat treatment (50, 60, 70, 80 °C) conditions. In conclusion, this study shows a high sensitivity, good selectivity and good stability of -HS, which can provide reference for the quality control and sulfur fumigation screening of GR.
Subject(s)
China , Chromatography, Liquid , Drugs, Chinese Herbal , Chemistry , Reference Standards , Fumigation , Gastrodia , Chemistry , Quality Control , Rhizome , Chemistry , Sulfites , Sulfur , Tandem Mass SpectrometryABSTRACT
To study the effect of different storage time on the chemical compositions and sulfur dioxide residues of sulfur-fumigated Gastrodiae Rhizoma (GR), and provide scientific basis for solving the quality and safety issues of sulfur-fumigated traditional Chinese medicinal materials. GR, sulfur-fumigated GR and its medicinal slices were stored under the same conditions, and then 8 active ingredients and sulfur dioxide residues were measured respectively. The results showed that the content of gastrodins in sulfur-fumigated GR and its medicinal slices was significantly lower than that in the non-fumigated GR. Moreover, the content of sulfur dioxide residue in sulfur-fumigated GR was significantly higher than that in its medicinal slices. That is to say, sulfur fumigation degree had significantly higher effect on GR quality as compared with its medicinal slices. During the whole storage time (8 months), the content of the eight chemical components in GR was not changed greatly in general. However, after the storage for 4 months, the content of 8 components and sulfur dioxide residues in all of GR samples were significantly changed. In particular, the content of sulfur dioxide residue in GR medicinal materials decreased up to 50% or more.
Subject(s)
Drug Storage , Drugs, Chinese Herbal , Chemistry , Fumigation , Gastrodia , Chemistry , Rhizome , Chemistry , Sulfur , Sulfur DioxideABSTRACT
Objective To evaluate the effect of endemic disease prevention in Shaanxi province caused by earthquake and supply a scientific basis for making decision regarding restoration and reconstruction after the earthquake.Methods Forty different drinking water samples were collected respectively from Mianxian county and Chencang district the earthquakes'most severely hit areas in Shaanxi province with drinking-water-borne fluorosis.The contents of fluorine and arsenic in the water was detected by Fluoride ion-selective electrode and hydride generation atomic fluorescence spectrometry,respectively.Two hundreds iodized salt and urine samples were collected from affected household women aged 18-45 in severely hit Ningqiang and Lveyang countie with iodine deficiency disorders.The contents of iodine in the salt was detected by semi-quantitative and of urinary iodine of household women was determined by AsⅢ-Ce4+ catalytic spectrophotometry.Two hundred children aged 7-12 were detected by clinical and X-ray examination from Ningqiang and Lveyang counties of Kashin-Beck disease prevailing areas.Hair and food samples from 20 children were collected.The contents of selenium were determined by 2,3-Diamino naphthalene fluorescence spectrophotometry and of T-2 toxin in the food was detected by ELISA.Results No incidents of serious hydrologieM or geological damage were found.Fluoride content in one of eighty water samples was beyond 1.5 mg/L,accounting for 1.25%;five samples were between 1.0 mg/L and 1.5 mg/L,accounting for 6.25%:the arsenic content of all samples were below 0.01 mg/L;all the salt samples from the home of household aged 18-45 were iodine salt,the median of urine iodine of household wonlen Was 303.03μg/L (21.40-1133.54 μg/L) and 325.21 μg/L(27.61 - 1191.46 μg,/L) in Ningqiang and Lveyang counties, respectively.No clinical and X-ray patients were found in the children aged 7 - 12, the average content of selenium in hair was (0.2638 ± 0.0875)mg/kg; the content of T-2 toxin in the forty food samples was below 100 μg/kg. Conclusions The prevention of the endemic disease in the four counties has been moderately affected by the earthquake, but the situation of endemic disease is stable and there is no evidence of the endemic disease.
ABSTRACT
<p><b>OBJECTIVE</b>To observe the impacts of doxapram on anesthetic efficacy and respiratory and circulatory functions during anesthesia with remifentanil given by target-controlled infusion (TCI) combined with propofol in painless artificial abortion.</p><p><b>METHODS</b>A total of 120 ASA I women requiring voluntarily painless artificial abortion were randomized into two equal groups. One group was given remifentanil by TCI at 2 ng/ml and propofol 1mg/kg (group I), and the other given remifentanil by TCI, propofol, and doxapram 0.6 mg/kg (group II). The anesthetic efficacy and respiratory and circulatory suppression during anesthesia were observed.</p><p><b>RESULTS</b>Both of the two groups showed satisfactory anesthetic efficacy with comparable the BIS values (P>0.05). No significant difference was noted in the induction time and awaking time between the two groups (P>0.05), but group I had more serious respiratory and circulatory depression than group II (P<0.05).</p><p><b>CONCLUSION</b>Doxapram may attenuate respiratory and circulatory depression during anesthesia with remifentanil given by TCI combined with propofol in painless artificial abortion, and provide comparable anesthetic efficacy.</p>