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Based on the data of 56 kinds of diseases and drug use in 100 kinds of cultivated Chinese herbal medicines, this paper used frequency analysis method to count the types of diseases and their drug use characteristics, and systematically analyzed the status of drug registration and monitoring standards for disease prevention and control of Chinese herbal medicines. The results showed that 14 diseases such as root rot, powdery mildew, and drooping disease were common in the production of Chinese herbal medicines. Among the 99 pesticides reported, 67.68% were chemically synthesized, 23.23% were biological pesticides, and 9.09% were mineral pesticides. Among the reported pesticides, 92.93% of them were low toxic, with relative safety. However, 70% of the production drugs were not registered in Chinese herbal medicines, and the phenomenon of overdose was serious. The current pesticide residue monitoring standards does not match well with production drugs in China. Although the matching degree between Maximum Residue Limit of Pesticide in Food Safety National Standard(GB 2763-2021) and production drugs is more than 50%, there are few varieties of Chinese herbal medicines covered. The matching degree between Chinese Pharmacopoeia(2020 edition), Green Industry Standard of Medicinal Plants and Preparations(WM/T2-2004), and production drugs is only 1.28%. It is suggested to speed up the research and registration of Chinese herbal medicine production and further improve the pesticide residue limit standard combined with the actual production, so as to promote the high-quality development of Chinese herbal medicine industry.
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Humanos , Agentes de Control Biológico , Medicamentos Herbarios Chinos , Residuos de Plaguicidas , PlaguicidasRESUMEN
Objective To investigate the health literacy level and health related behavior of high school students in Xuhui District of Shanghai, and to provide basis for health literacy intervention. Methods Using qualitative and quantitative methods, the quantitative part adopted the multi-stage stratified sampling method.A total of 355 aged 15-18 from senior high schools in Xuhui District were selected for face-to-face survey.The qualitative part adopted the form of group interviews, and 20 students from two senior high schools in Xuhui District were selected for interviews. Results The health literacy level of high school students in Xuhui District was 27.32%, which was lower than that of adults in the same period.Among the three dimensions, the level of basic knowledge and concept literacy was the highest, followed by the level of health basic skills literacy, the level of healthy lifestyle and behavior literacy was the lowest; among the six types of health problems, the level of infectious disease prevention literacy and basic medical literacy was at a low level.Statistical analysis of the correct answer rate showed that the answer rate of various questions in the health literacy survey was quite different.Further analysis of the health-related behavioral characteristics of high school students showed that most students had correct health concept and awareness of health change, but they encountered resistance in the process of knowledge transformation into action, which mainly came from academic burden, school education, peer influence and so on.The main channels for high school students to obtain health-related information were classroom, parents, school and various media.They had a high degree of trust in medical professionals and encountered difficulties in seeking health information. Conclusion The health literacy level of senior high school students in Xuhui District of Shanghai needs to be improved.We should explore a health intervention model more suitable for the behavioral characteristics of this group weak links in their health literacy.
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The study aims at investigating the grade standard and the quality characteristic of Pinelliae Rhizoma for commodity specification, which provides the reference for its grade standard formulation. 42 Pinelliae Rhizoma simples were collected from 5 medicinal materials markets and 2 producing areas. Based on the previous herbalogical study and market investigation, we combined with the data analysis to select the grading indicators using SPSS software for descriptive statistical analysis, analysis of variance, K-cluster analysis and correlation analysis. According to the actual production condition, we developed the grading standards of Pinelliae Rhizoma. Moreover, we compared the internal indicators(water, total ash, leachate and guanosine) of Pinelliae Rhizoma at various grade levels, and analyzed the correlation between appearance traits and internal indicators. The herbalogical study and market research found that the Pinelliae Rhizoma was better in large, solid and white. The results from descriptive and variance analysis showed that the appearance of Pinelliae Rhizoma was significantly different in weight per grain and grain number of 500 g. Referring to the 2015 Chinese Pharmacopoeia and the production practice, we use the length, weight per grain and grain number of 500 g as the classification index of Pinelliae Rhizoma. The results from correlation analysis showed that there was no significant correlation between the appearance of Pinelliae Rhizoma and the intrinsic quality index. In addition, we found there was no significant difference in the content of the intrinsic index except for the total ash and the extract. The current study established the classification index of the product specification and grade standard of Pinelliae Rhizoma with length, weight per grain and grain number of 500 g as the index, which can provide the basis for the classification of the product specification and grade of Pinelliae Rhizoma market.
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Medicamentos Herbarios Chinos , Estándares de Referencia , Pinellia , Química , Rizoma , QuímicaRESUMEN
Through the records of herbal and market investigations,the samples collected from different market and origin were analyzed,and the appearance character indexes were determined in order to revise the commodity specification and grade standard of Dipsaci Radix combined with production practice. This is also to analyze association of the appearance with quality different of intrinsic components. The investigation results indicated the root's long and thick was better,and atrovirens of fracture surface,there are most of the market is dominated by the ungraded goods. Through principal component analysis,variance analysis and cluster analysis,combining with the feasibility of actual operation and herbal record,the length,middle diameter and phloem color were filtrated and the specification was divided into 2 ranks: the selection and ungraded goods,and the grade was divided into 2 ranks for the selection goods: big,small selected goods. Moreover,there were no significant correlation between the appearance and the intrinsic quality index of Dipsaci Radix,the content of extractum were significantly positive correlation with the moisture,but had extremely negative correlation with the total ash. Multiple comparisons indicated that the content of the moisture,extractum and asperosaponin VI of the selected goods were higher than the ungraded goods,while the total ash content were lower,and they hasn't significant difference. The commodity specification and grade standard of Dipsaci Radix as a basis provide commodity specification and grade standard of communities and standardizing market trade order.
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Análisis por Conglomerados , Dipsacaceae , Química , Medicamentos Herbarios Chinos , Estándares de Referencia , Raíces de Plantas , Química , Plantas Medicinales , Química , Análisis de Componente PrincipalRESUMEN
This paper is aimed to study the commodity specification and grade standard of Galla Chinensis,for standardizing market order and guide the market circulation,and provide a basis for standardization of Galla Chinensis. With 33 samples of Galla Chinensis from market as the object of the research,the herbal textual research and market research were carried out. Then the grading indicator were selected by the descriptive statistics,principal component analysis and cluster analysis,combining with production practice,the commodity specification and grade standard of Galla Chinensis were set out. The data of moisture,ash,gallic acid as the internal index were used to analyze the relationship between the quality difference between grades and the appearance characters and the intrinsic composition. Herbal textual research and market research found that the high quality of Galla Chinensis characterized with large,complete,wall thick,grayish brown characteristics,and Galla Chinensis could be divided into gallnuts and horned gall. Through principal component analysis and cluster analysis,combining actual production and herbal record,screening,the length,diameter,single weight,the number of 500 g were determined as 4 grading indicators,the commodity specification was divided into two: gallnuts and horned gall,the grades was divided into two: selected goods and mixed goods. The result of correlation analysis showed there was no significant correlation between the internal index and the appearance characters of Galla Chinensis. The result of multiple comparison showed that the ash content of the selected goods was smaller than that of the mixed goods,but it did not reach significant. The content of gallic acid of the selected goods and the mixed goods of gallnuts were higher than selected goods of horned gall,and higher than mixed goods of horned galls. Using the length,diameter,single weight,the number of 500 g as the appearance characters index. Combining with the herbal textual research and market research,we have divided two specifications and two grades for the commodity specification and grade standard of Galla Chinensis,which can provide a basis for industry standards and national standards.
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Medicamentos Herbarios Chinos , Estándares de Referencia , Ácido Gálico , Tumores de Planta , Control de CalidadRESUMEN
<p><b>OBJECTIVE</b>To investigate the effect of propofol and operative trauma on the neurodevelopment and cognitive function of the developing brain and its mechanism.</p><p><b>METHODS</b>A total of 104 postnatal day 13 Sprague-Dawley rats were randomly divided into 4 groups: control group (treated by 7.5 mL/kg saline and sham surgery), propofol group (treated by 75 mg/kg propofol), surgery group (with abdominal surgery under local anesthesia) and propofol+surgery group (with abdominal surgery under local anesthesia plus 75 mg/kg propofol anesthesia). Thirteen rats from each group were randomly selected for detecting the content of TNF-α in the hippocampus and the expression levels of caspase-3 and c-fos in the brain. Morris Water Maze test was used to detect the cognitive ability of the other rats at 60 days old, after which TNF-α content in the hippocampus and caspase-3 and c-fos expressions in the brain were detected.</p><p><b>RESULTS</b>In 13 day-old rats, TNF-α level and caspase-3 and c-fos expressions differed significantly between the surgery group and the other 3 groups (P<0.05) and were similar among the control group, propofol group and propofol+surgery group (P>0.05). In 60-day-old rats, Morris water maze test results, TNF-α level or expressions of caspase-3 and c-fos showed no significant differences among the 4 groups.</p><p><b>CONCLUSION</b>Abdominal surgery can induce inflammation in the hippocampus and neuroapoptosis in neonatal rats rather than adult rats. Single-dose propofol anesthesia does not significantly affect neurodevelopment of young rats, and can relieve central inflammatory reaction induced by surgical trauma.</p>
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Objective To measure the catalytic effect of ammonium salt solution on hydrogen production , and analyze the relationships between hydrogen concentration and oxygen concentration , oxidation-reduction potential , and pH in solution . Methods Magnesium and water reacted at different concentrations of NH4Cl, (NH4)2SO4, NH4HCO3,(NH4)2CO3, NaHCO3 and Na2SO3 in 40℃water bath for 0, 2, 4, 6, 8 and 10 hours, and above-mentioned four indicators of ammonium salt solution were measured after these reactions .Results The hydrogen concentration in the solution increased with the reaction time and the concentration of the ammonium salt solution .Oxygen concentration and oxidation-reduction potential decreased .The hydrogen concentration in the solution was significantly negatively correlated with the oxygen concentration (r=-0.984).pH increased with the hydrogen concentration in the ammonium salt solution .Conclusion The ammonium salt solution has a good catalytic effect on magnesium and water reaction .NH4Cl has the strongest catalytic effect under the same reaction conditions,followed by (NH4)2SO4, NH4HCO3 and (NH4)2CO3.Thus, this study can provide detailed data on hydrogen production in different solutions .
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The study aims at evaluating genetic diversity and medicinal quality of cultivated germplasm in Rehmannia glutinosa, and providing theoretical guidance for screening excellent germplasm. The genetic diversity of 21 species of R. glutinosa were analyzed by SRAP molecular markers, and the catalpol and verbascoside was determined by HPLC. The mass fraction of catalpol and verbascoside in R. glutinosa germplasm were respectively in the range of 2.393%-6.519% and 0.063%-0.478%, the germplasm 14, 16, 15 and 20 germplasm, witch catalpol and verbascoside content was higher. A total of 57 bands were produced by 10 primer, among which 40 polymorphic bands were polymorphic bands, and the percentage of polymorphic loci was 8.77%-54.39%, the Nei's genetic diversity index (H) was 0.374 1, Shannon's polymorphism information index (I) was 0.546 6. Gst and gene flow Nm were 0.608 8 and 0.321 3, respectively. Based on the genetic uniformity, 21 species of germplasm were grouped into 2 categories. The genetic diversity level of R. glutinosa was medium low. The comprehensive consideration of the genetic diversity and the content inculde catalpol and verbascoside, germplasm 7 and germplasm 18 could be used as the preferred materials for the cultivation of reticulum. Germplasm 15 and 16 can be used as the preservation and breeding object of rhubarb germplasm.
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Animales , Flujo Génico , Variación Genética , Filogenia , Fitomejoramiento , Plantas Medicinales , Genética , Rehmannia , GenéticaRESUMEN
Objective To investigate the hydrogen production rate of hydrogen rods in different solute solutions ,and to study the correlations between hydrogen and oxygen concentrations in the solution and the relationships between hydrogen and oxidation-reduction potential .Methods The hydrogen rich solution was produced using metal magnesium to react with water.The experiment was divided into NaCl , Na2SO3, Na2SO4, CH3COOH, and CH3COONa solution groups, respectively, with 0%, 0.2%, 0.9%and 3%in different concentration solutions .The hydrogen content of the corresponding solution was determined at 0, 2, 4, 6, 8, and 10 h.Moreover, the oxygen content in the solution , redox potential and pH value were simultaneously determined .Results Within the same reaction time , the higher concentration of NaCl , the higher the hydrogen production rate .The amount of hydrogen in the solution was negatively correlated with that of oxygen ( R2 =0.9306).The higher the hydrogen content, the lower the oxygen content.With the amount of hydrogen in the solution increasing, oxidation was reduced while pH was increased.The hydrogen-producing rate of hydrogen rods in different solute solutions was the highest with Na2SO3,followed by NaCl and Na2SO4(P<0.05 or P<0.01).In the acetic acid solution, the hydrogen-producing rate of rods was significantly higher than in the sodium salt solution (P<0.05).The hydrogen content increased gradually with time .The oxygen concentration was significantly lower than in the sodium salt solution ( P <0.05), and the oxygen content decreased over time .Conclusion The type and concentration of solutes in solutions and the oxygen content and acidity of a solution have significant influence on the hydrogen -producing rate of hydrogen rods. Therefore, by adjusting the type and concentration of solutes in the solution, we can prepare different types of hydrogen concentration solutions , which can provide detailed reference parameters for hydrogen production of hydrogen rods in clini -cal and practical applications.
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BACKGROUND: The whole herb of Lagochilus ilicifolius has been used as a folk medicine for treating hemostatic, inflammation and ulcer in China. There were only limited reports on its chemical constituents, and no reports on its pharmacology study. OBJECTIVE: To isolate compounds from the whole herb of L. ilicifolius and evaluate their cytotoxic activity. MATERIALS AND METHODS: The column chromatographic techniques were used for separating the constituents of the n-butanol-soluble fraction of the 95% ethanol extract from the whole plant of L. ilicifolius. The structures of one new lignan and two known lignans were elucidated on the basis of spectroscopic analyses and comparison with literature data. The cytotoxic activities of these three lignans were evaluated using the MTT-assay against PC12 cell line derived from rat adrenal pheochromocytoma. RESULTS: The new lignan was identified as erythro-1-[(4-O-ß-D-glucopyranosyl-3-methoxyl)-phenyl]-2-[(5'-methoxyl)-pinoresinol]-propane-1,3-diol (1), and two known lignans were identified as tortoside C (2) and sisymbrifolin (3). The new lignan exhibited significant cytotoxic activity against PC12 cell line with IC50 value of 1.22 ± 0.03 µmol/L. CONCLUSIONS: A new lignan, erythro-1-[(4-O-ß-D-glucopyranosyl-3-methoxyl)-phenyl]-2-[(5'-methoxyl)-pinoresinol]-propane-1,3-diol and two known lignans were isolated from the whole herbs of L. ilicifolius. The two known lignans were reported for the first time in the genus Lagochilus. Three lignans were evaluated for in vitro cytotoxic activity. The new lignan showed relatively strong cytotoxicity against PC12 cell line, while sisymbrifolin and tortoside C exibited no cytotoxicity.
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Objective: To study the chemical constituents from the whole herb of Lagochilus platyacanthus. Methods: The chemical constituents were isolated and purified by various chromatographic techniques and the structures were identified by spectral analysis. Results: Twenty-one compounds were isolated from the 95% ethanol extract of L. platyacanthus, including 15 flavonoids: apigenin-7, 4'-dimehyl ether (1), acacetin (2), apigenin (3), luteolin-7, 3', 4'-trimethyl ether (4), luteolin-7, 4'-dimethyl ether (5), diosmetin (6), chrysoeriol (7), quercetin-3-O-rutinoside-7-O-glucoside (8), rutin (9), horridin (10), apigenin-6, 8-di-C-β-D-glucopyranoside (11), isorhamnetin-3-O-rutinoside (12), isorhamnetin-3-O-robinobioside (13), isorhamnetin-3-O-β-D-glucoside (14), and isorhamnetin-3-O- rutinoside-4'-O-glucoside (15); three lignans: 1-(4-hydroxy-3-methoxy)-phenyl-2-[4-(1, 2, 3-trihydroxypropyl)-2-methoxy]-phenoxy- 1, 3-propandiol (16), (+)-isolarisiresinol 3-α-O-β-D-glucopyranoside (17), and (-)-isolarisiresinol 3-α-O-β-D-glucopyranoside (18); two iridoids: 8-O-acetylharpagide (19) and geniposidic acid (20), and one phenylethanoid glycoside: lavandulifolioside (21). Conclusion: All the compounds are obtained from this plant for the first time. The compounds are isolated from the plants in genus Lagochilus Bunge for the first time except compounds 1, 3, 9, 19, and 20.
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<p><b>OBJECTIVE</b>To explore the therapeutic effects of hemoperfusion (HP) with continuous venovenous hemofiltration (CVVH) on the patients with acute paraquat poisoning.</p><p><b>METHODS</b>Ninety-one patients with acute paraquat poisoning were randomly divided into HP group (49 cases) and HP-CVVH group (42 cases). The mortality, survival duration and the death causes between the two groups were compared and analyzed.</p><p><b>RESULTS</b>There were no significant differences in mortality (59.2% versus 61.9%) between the two groups. The mean time between poisoning and death in HP-CVVH group was (4.9 +/- 3.1) days, which was significantly longer than that (3.5 +/- 2.0) days in HP group (P < 0.05). The death proportion on 4th day after poisoning in HP group was 62.1% (18/29), which was significantly higher than that (30.8%, 8/26) in HP-CVVH group (P < 0.05). The hypoxia appeared in 4.3 +/- 2.5 days after poisoning in HP-CVVH group, which was significantly longer than that (3.2 +/- 1.9) days in HP group (P < 0.05). The mortality due to respiratory failure in HP group was 20.4% (10/49), which was significantly lower than that (40.5%, 17/42) in HP-CVVH group (P < 0.05). The incidence of acute renal failure in HP group was 63.3% (31/49), which was significantly higher than that (40.5%, 17/42) in HP-CVVH group (P < 0.05).</p><p><b>CONCLUSION</b>The combined therapy of HP and CVVH can prevent the patients with acute paraquat poisoning from early death and prolong the survival duration, but can not reduce mortality for the patients with acute paraquat poisoning.</p>
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Adolescente , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Adulto Joven , Hemofiltración , Hemoperfusión , Paraquat , Intoxicación , Intoxicación , Mortalidad , Terapéutica , Resultado del TratamientoRESUMEN
<p><b>OBJECTIVE</b>To study the correlation factors of acute paraquat intoxication prognosis.</p><p><b>METHODS</b>The early paraquat concentration in plasma and urine, leukocyte count, hepatic and renal function, amylase, electrolyte and the parameters of arterial blood gas were analyzed retrospectively in 111 patients with acute paraquat intoxication.</p><p><b>RESULTS</b>43 cases (38.7%) of all the 111 patients survived and the other 68 cases (61.3%) died. The patient, whose paraquat concentration was not more than 8.0 µg/ml in plasma and 276.0 µg/ml in urine, could survive. But some patients could die, only if there was no paraquat found in plasma. The paraquat levels in plasma and urine were significantly lower in survivors [(0.82 ± 1.70), (28.12 ± 51.17) µg/ml] than in nonsurvivors [(9.32 ± 12.04), (384.53 ± 597.93) µg/ml, respectively] (P < 0.01). The levels of leukocyte count, serum creatinine, aspartate aminotransferase (AST), and amylase were significantly higher in nonsurvivors than in survivors (P < 0.05, P < 0.01). In addition, metabolic acidosis was easier to appear in nonsurvivors. The multiple logistic regression analysis indicated that the paraquat concentration in plasma and urine, leukocyte count, creatinine and base excess were all related to survival.</p><p><b>CONCLUSION</b>The higher paraquat concentration in plasma and urine, leucocytosis, renal dysfunction and metabolic acidosis are all important factors for the prognosis of paraquat intoxication.</p>
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Adolescente , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Adulto Joven , Acidosis , Enfermedades Renales , Leucocitosis , Paraquat , Sangre , Intoxicación , Orina , Pronóstico , Estudios RetrospectivosRESUMEN
<p><b>OBJECTIVE</b>To develop a suitable hypoxic injury model, which is important for revealing pathological molecular mechanism of hypoxia.</p><p><b>METHODS</b>We focused on C. elegans by treatment with different hypoxic times and systematically observed mortality, movement, Cellular morphology and the related-protein expression of the animals.</p><p><b>RESULTS</b>We demonstrated that hypoxia (0.2% partial pressure of oxygen) induced morphological cell defects, and then leading to death of C. elegans. The mortality of C. elegans increased along with hypoxic time, while hypoxia-inducible factor (HIF-1) was significantly up-regulated. In addition, by using neuron-specific transgenic wonns with green fluorescent protein--we observed the neuron-specffic injury caused by hypoxic stress.</p><p><b>CONCLUSION</b>We successfully established an effective, convenient physical hypoxic model of C. elegans, which will facilitate the studies of hypoxic pathology and molecular mechanisms of hypoxic response in the future.</p>
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Animales , Caenorhabditis elegans , Fisiología , Proteínas de Caenorhabditis elegans , Metabolismo , Modelos Animales de Enfermedad , Hipoxia , Factor 1 Inducible por Hipoxia , Metabolismo , Neuronas , Patología , Factores de Transcripción , MetabolismoRESUMEN
We assessed the potential eco-toxicological risks of the herbicide acetochlor on fungal communities in the microcosm of black soil using 28S rRNA gene-PCR-DGGE and clone library analysis. The acetochlor was applied to black soil at four concentrations (0-control, 50, 150, and 250 mg/kg). The DGGE fingerprint patterns indicated that acetochlor stimulated fungal communities at day 7 after application, after which there was a suppression effect. The fungal communities in acetochlor-treated soil gradually became more like that of the control during the 60-day experimental period. Diversity indices in the 50 and 150 mg/kg acetochlor treatments changed more rapidly than in the 250 mg/kg acetochlor treatment. The cluster analysis indicated a significant change in fungal community structure after application of acetochlor. The impacts were markedly greater in the 150 and 250 mg/kg acetochlor treatments compared with the 50 mg/kg acetochlor treatment. Sequencing of clones showed that acetochlor application resulted in an increase in pathogenic and non-cultivatable fungal populations, which could increase the risk of plant disease outbreaks.
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Hongos/efectos de los fármacos , Herbicidas/farmacología , Microbiología del Suelo , Contaminantes del Suelo/farmacología , Toluidinas/farmacología , Hongos/genética , Hongos/crecimiento & desarrollo , Biblioteca de Genes , Datos de Secuencia Molecular , Filogenia , Reacción en Cadena de la Polimerasa , ARN Ribosómico 28S/genéticaRESUMEN
Objective To explore a sensitive method and index to evaluate renal functional reserve (RFR) in patients with early diabetic nephropathy (DN) using protein loading-scintirenography.Methods Fifty subjects were studied and divided into 3 groups.Group one (G1) consisted of 14 healthy volunteers;Group two (G2) consisted of 15 patients with type 2 diabetes mellitus (DM) and normoalbuminuria; Group three (G3) consisted of 21 patients with type 2 DM and microalbuminuria.All subjects underwent baseline and protein loading-99 Tcm-DTPA scintirenography within one week.RFR was calculated as the difference between stimulated and baseline glomerular filtration rate (GFR), time of peak ( Tb ), time of half excretion ( C1/2 ), residual rate at 20 min ( C20/b ) .Variance analysis and t-test were used to analyze the group differences.Results ( 1 ) The RFR in terms of GFR had statistical difference between any two groups (t=14.884, 32.180, 16.042, all P<0.01).After protein-loading, the GFR of G1, G2 and G3 increased 20.1, 10.9 and 2.2 ml·min-1·1.73 m-2 respectively.Therefore, the RFR decreased before microalbuminuria in type 2 DM patients.(2)There was statistical difference between the RFR of G1 and G2 in terms of C1/2 (t = 5.505, P<0.05 ), and between G1 and G3 ( t = 8.914, P<0.01 ).(3) There was statistical difference of the RFR in terms of TP between G1 and G3 (t = 5.690, P < 0.01 ).(4) There was statistical difference of the RFR in terms of C20/b between G1 and G3 (t= 4.376, P<0.05 ).Conclusions 99Tcm-DTPA protein loading-scintirenography is an effective method for measuring RFR to evaluate early DN in type 2 DM patients.
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Heterogeneous nuclear ribonulcleoprotein(hnRNP)plays a variety of roles in pre-mRNA splicing, nuclear export of mRNA and turnover. It is reported that hnRNP not only takes part in regulating the development of neuron and glial cells, but is closely related to various central nervous system diseases. This paper reviewed the structure, function and the role of hnRNP in the central nervous system in order to provide new insight into the molecule mechanism of nervous system diseases.
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The whole sec2 DNA fragment was obtained by polymerase chain reaction with total genomic DNA extracted from Staphylococcus aureus. The newly discovered gene contains 717 bp (GenBank Accession number AY450554) and encodes 239 amino acids in accordance with the protein sequence reported in GenBank. The sec2 gene was fused to anti-Her-2 scFv gene with a DNA linker at the upstream to construct the fusion gene of immunotoxin which was subcloned to expression vector pET-32a(+) and expressed efficiently in Escherichia coli. The purified fusion immunotoxin could target the HER-2 overexpressed by breast tumor cell SK-Br-3 in vitro and demonstrate tumor-inhibition effect on SK-Br-3.
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Enterotoxinas/metabolismo , Escherichia coli/genética , Inmunotoxinas/genética , Inmunotoxinas/inmunología , Receptor ErbB-2/inmunología , Antineoplásicos , Línea Celular Tumoral , Enterotoxinas/genética , Expresión Génica , Humanos , Inmunotoxinas/metabolismo , Datos de Secuencia Molecular , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismoRESUMEN
Neuroglobin(NGB),widely and specially expressed in neurons of various vertebrates,was reported to be a scavenger of reactive oxygen species and/or a stress-responsive sensor for neuroprotection of hypoxic and ischemic insults.However,the underlying mechanism remained unknown.It is important for the functional study of NGB by preparing the protein samples.To address it,the human neuroglobin cDNA fragment was amplified by RT-PCR from human fetus brain and cloned into the prokaryotic expression vector pBV220,and then transformed into E.coli HB101 cells for expression.The expressed protein was purified by gel filtration and anion exchange column followed with SDS-PAGE and Western blot identification,and then desalting by sephadex G-25 medium.The prepared neuroglobin was further identified by mass spectrometry and N-terminal amino acid sequencing analysis.The expressed bacterium,the lysate supernatant and the purified protein samples had visible red color,showing a typical activity of the globin family proteins.In conclusion,the neuroglobin was not only expressed in soluble form with high-efficiency in E.coli,but could also be easily purified with only two steps.The preparation of the NGB proteins will advance the neuroprotective function and mechanism studies of the novel globin.
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DNA mismatch repair gene mutS (2.56 kb) was PCR modified and cloned into a secretive prokaryotic expression vector pET32a (+) which carries a N-terminal His.tag + and thioredoxin sequence. MutS protein was expressed with high level after IPTG induction using the strain E. coli AD494(DE3). SDS-PAGE revealed that the expected protein with a molecular weight of 108 kD which is about 35% of the total bacterial proteins is almost soluble. The expected protein was purified directly by immobilized metal (Ni2+) chelation affinity chromatography and the purity is over 90%. MutS protein activity verified using mismatch DNA showed that the expression product can recognize and bind to base-pair mismatch specifically.