Technological improvement for microorgnism isolation is important since isolation provides substantial materials for the exploitation of new microbial resources. In this study, a new approach, dispersion and differential cetrifugation (DDC), was applied in the isolation of acidophilic and acidoduric streptomycetes from 12 acid soil samples. Contrast with traditional method, the new approach yielded satisfying results with 2 - 20 times isolation efficiency and good selectivity. 45 representatives out of 249 streptomycetes isolates, which belonged to 12 color groups, showed morphology and cell wall type consistent with streptomycetes. The optimum pH range for their growth were between pH 4.5 - 5.5. It is proved that we succeeded in the rare-streptomycetes isolation using DDC approach.

Background and purpose:Chemotherapy plays an important role in the treatment of breast carcinoma by inhibiting the tumor growth and inducing the apoptosis.MAPK transduction pathway is closely related to proliferation and apoptosis of varieties of tumor cells,inhibition of MAPK pathway may increase the efficiency and decrease the toxicity of chemotherapy.Our study was to investigate the effect of MEK inhibitor PD98059 in response of breast cancer cell lines to Epirubicin.Methods:Human breast cancer cell lines MCF-7 and MCF-7/ADR were used as cell models.Epirubicin(EADM),PD98059(inhibitor of MAPK Kinase-MEK),or EADM+PD98059 was added into the culture medium,the expression of MEK2 and p-ERK were measured by Western blot,the growth of the two cell lines were measured by MTT.Results:ERK activity was elevated in MCF-7 after the treatment of EADM,the cells were more sensitive to EADM if combined with PD98059,while in MCF-7/ADR,ERK activity kept unchanged after EADM treatment,and PD98059 has no effect on the sensitivity of cells to EADM.Conclusion:MAPK signal transduction may be activated in some cells treated by EADM,adding inhibitor of MAPK signal transduction could improve the sensitivity of the cells to EADM.

The GFP(green fluorescence protein)-streptavidin(SA) bi-functional fusion protein was generated and characterized in order to demonstrate novel platform for efficiently and durably modifying the cell surface with SA-tagged bi-functional proteins.The GFP-SA/pET24d construct was generated and expressed in BL21(DE3) host bacteria at the high level.The recombinant protein GFP-SA was purified through the Ni-NTA affinity chromatography,and then refolded.After biotinylation B16 tumor cells were modified with GFP-SA bi-functional fusion protein and then subjected to fluorescent microscopy and FACS analysis.The effect of surface modification on the viability and growth of B16.F10 tumor cells was evaluated by MTT staining.The GFP-SA recombinant fusion protein was expressed in BL21(DE3) at about 20 % of total bacterial proteins.The GFP-SA bi-functional fusion protein exhibited the bi-functionality,i.e.,SA-mediated high-affinity binding to biotinylated cell surfaces and GFP-emitted green fluorescence.The cell surface modification with GFP-SA bi-functional fusion protein did not affect the viability and growth of the modified B16.F10 tumor cells significantly.The GFP-SA bi-functional fusion protein was obtained and could be displayed efficiently on the surface of the biotinylated B16.F10 tumor cells through the specific and tight interaction between streptavidin and biotin,thus can be used as good trace protein and experimental control in the development of surface-modified tumor vaccine.

By reverse transcription-polymerase chain reaction (RT-PCR), an open reading frame of pathogenesis-related protein 1 (PR1) was isolated from Panax notoginseng and named as PnPR1. Molecular and bioinformatic analyses of PnPR1 revealed that an open reading frame of 501 bp was predicted to encode a 166-amino acid protein with a deduced molecular mass of 18.1 kD. Homology analysis showed that the deduced amino acid sequence of PR1 protein of Panax notoginseng had a high similarity with other higher plants had the same conservative structure domain of cysteine-rich secretory protein (CAP). The recombinant expressed plasmid pET28a(+)-PnPR1 was expressed in Escherichia coli BL21. The expression conditions were optimized by induction at different times, different temperatures, different IPTG concentrations and different giving times. The optimum expression condition was 0.4 mmol.L-1 IPTG at 28 degrees C for 20 h. The successful expression of PnPR1 provides some basis for protein purification and preparation of the monoclonal antibody.
Amino Acid Sequence ; Cloning, Molecular ; Escherichia coli ; metabolism ; Molecular Weight ; Open Reading Frames ; genetics ; Panax notoginseng ; chemistry ; Phylogeny ; Plant Proteins ; genetics ; metabolism ; Plants, Medicinal ; chemistry ; Reverse Transcriptase Polymerase Chain Reaction ; Sequence Alignment

Objective To investigate the possible role of interstitial cell of Cajal (ICE)in the pathogenesis of diabetic gastroparesis and the protective effect of"Xiaopi prescription (消痞方)".Methods Fifty healthy male Spregue-Dawley (SD) rats were randomly divided into 5 groups (each n=10):normal, model,and 3 Xiaopi prescription groups:low,middle and high dosages.Diabetes was induced by intravenous injection of alloxan,and equal amount of normal saline was intravenously injected in the normal group.Gastric lavage method was used to administer the traditional Chinese medicine decoction of Xiaopi prescription in corresponding amount (5,10 and 20 g?kg~(-1)?d~(-1)) in respective low,middle and high dosage groups.In the normal control group and diabetic model group,only equal amount of normal saline was administered into the stomach.Gastric emptying rate was measured by method of nutritious semisolid paste;c-kit positive cells of ICC were quantitatively measured with immunohistochemistry assay and computer image analysis system;c-kit mRNA positive cells were quantitatively measured with in situ hybridization and computer image analysis system.Results①Gastric emptying rate:The rate was significantly lower in the model group than that in the normal control group (P0.05),but higher than those in the model group and the low dosage group (all P0.05).②c-kit immmunohistochemistry:c-kit positive cell presented yellow in color,and its membrane was stained yellow,this kind of cells primarily were distributed around the neural plexus in the inter-space between the circular and longitudinal muscular fibers, and around the ganglionic cells forming"sheath-like"structure.The results of numbers of c-kit positive cells in the various groups:the number of the cells in the model group was significantly lower than that in the normal group (P0.05),but the numbers in the former two dosage groups were obviously higher than those in the model group (all P0.05),being significantly lower than that in the normal group,middle and high dosage groups (all P0.05),but obviously higher than those in the model group (all P0.05),being significantly lower than that in the normal,middle and high dosage groups (all P

OBJECTIVETo provide new germplasm materials for breeding new varieties of Pseudostellaria heterophylla.

METHODThe method of single plant selection was adopted, with the comparative experiments being carried out under the same conditions in Shibing county. The 8 plants of Shibing SB-4 were compared respectively with factor analysis for 27 phenotypic traits and 8 yield traits, and single factor variance analysis for the contents of polysaccharides.

RESULTUsing factor analysis, 27 phenotypic traits were classified into 7 principal divisors and 8 yield traits were simplified into 3 principal divisors. The 4 strains of P. heterophylla, ZT-01, ZT-02, ZT-06 and ZT-07, performed better than others in the phenotypic traits, and ZT-01, ZT-02, ZT-03 and ZT-07 in the yield traits. The contents of polysaccharides of ZT-01, ZT-02, ZT-05 and ZT-08 showed significantly higher value.

CONCLUSIONThere is significant difference among the 8 strains of P. heterophylla in phenotypic traits, yield traits and quality traits, making it possible to select certain strains for different purposes. ZT-01 and ZT-02 can be breaded further. ZT-06 and ZT-07 were used as ornamental cultivars for its great phenotypic traits. ZT-03 with good resistance and high yield was taken as resistant variety, and ZT-05 would face next selection on the basis of its high content of polysaccharide.

Breeding ; Caryophyllaceae ; chemistry ; growth & development ; China ; Phenotype ; Polysaccharides ; analysis

OBJECTIVETo evaluate the value of dual source computed tomography coronary angiography (DSCT-CA) on detecting in-stent restenosis (> 50% luminal narrowing) in symptomatic patients referred for quantitative coronary angiography (QAC).

METHODSFifty five patients (43 males) with chest pain after coronary stent implantation within 6 - 12 months were evaluated by DSCT-CA and QAC. The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of DSCT-CA were calculated using coronary angiography as gold standard.

RESULTSEighty nine stents were implanted. In-stent restenosis was evidenced in 28 stents (31.5%) by QAC. The sensitivity, specificity PPV and NPV of DSCT-CA for the diagnosis of in-stent restenosis was 89%, 87%, 76% and 95%, respectively. Diagnostic efficiency was not affected by heart rate and the sensitivity was 0.94 vs. 0.82, the specificity 0.88 vs. 0.90, the PPV 0.76 vs. 0.75 and the NPV 0.97 vs. 0.93 (all P > 0.05) between patients with heart rate < 70 beats/min and patients with heart rate ≥ 70 beats/min. The sensitivity (84% vs. 100%), specificity (81% vs. 96%), PPV (70% vs. 90%) and NPV (91% vs. 100%) were similar between overlapping or bifurcations stents and single stents. The specificity (100% vs. 80% vs. 66%) and PPV (100% vs. 95% vs. 53%) were significantly higher in the groups with stents ≥ 3.50 mm, stents 3.00 mm than in stents ≤ 2.75 mm (both P < 0.05).

CONCLUSIONDiagnostic efficiency of in-stent restenosis with DSCT-CA in the large diameter stent is better than in the small diameter stent and the diagnosis efficacy is not affected by heart rate and stent distribution.

Adult ; Aged ; Coronary Angiography ; methods ; Coronary Restenosis ; diagnostic imaging ; Female ; Graft Occlusion, Vascular ; diagnostic imaging ; Humans ; Male ; Middle Aged ; Predictive Value of Tests ; Prospective Studies ; Sensitivity and Specificity ; Stents ; Tomography, X-Ray Computed ; methods

Animals ; Antibodies, Monoclonal ; immunology ; Atrazine ; analysis ; immunology ; Chromatography, Gas ; Chromatography, High Pressure Liquid ; Enzyme-Linked Immunosorbent Assay ; Food Contamination ; analysis ; Herbicides ; analysis ; immunology ; Mass Spectrometry

The infection status of anisakid larvae was examined in 290 marine fish of 25 species and in 108 cephalopods of 3 species purchased in Bayuquan region, Yingko city nearby the coast of the Bohai Sea from may to August 1992. A total of 7,327 larvae were collected from 156 fish of 19 species and 8 squids of one species. The 3rd-stage larvae of Anisakis simplex were collected from 121 fish (63.4%) of 15 species (N = 191) and from 8 squids (14.8%) of one species (N = 54), and they were total, 5,992 (81.8%). Out of remaining 1,335 larvae, 154 (2.1%) were classified as Thynnascaris type B from 23 fish of 4 species, 1,013 (13.8%) as Thynnascaris type C from 79 fish of 13 species. 164 (2.2%) as Hysterothylacium China type V from 20 fish of 4 species, 3 (0.04%) as Raphidascaris from 3 fish of 2 species and one was Pseudoterranova decipiens larva.
Animal ; Anisakiasis/veterinary* ; Anisakiasis/parasitology ; Anisakiasis/epidemiology ; Anisakis/isolation & purification ; Anisakis/classification* ; China ; Fish Diseases/parasitology* ; Fish Diseases/epidemiology ; Fishes ; Larva ; Seawater ; Squid/parasitology*

OBJECTIVETo study the purification, refolding and bioactivity of streptavidin-tagged human tumor necrosis factor-alpha (SA-TNF-alpha) bi-functional fusion protein.

METHODSSA-TNF-alpha fusion protein was expressed in BL21(DE3) host bacteria, purified using Ni-NTA affinity chromatography and refolded by dilution and dialysis followed by identification using Western blotting. The effect of SA-TNF-alpha fusion protein against L929 cells was evaluated by MTT assay. Flow cytometry was used to analyze the surface modification of biotinylated MB49 tumor cells by SA-TNF-alpha fusion protein.

RESULTSRecombinant SA- TNF-alpha fusion protein was expressed in BL21(DE3) at about 30% of total bacterial protein, with a purity of about 95% after purification. The SA-TNF-alpha fusion protein existed as dimmers, tetramers and higher order structures after refolding. The fusion protein exhibited a bi-functionality by inhibiting L929 cells and SA-mediated high-affinity binding to biotinylated cell surfaces, with an anchor modification rate of above 90%.

CONCLUSIONThe dimmers, tetramers and higher order structures of the obtained SA-TNF-alpha fusion protein all exhibit a bi-functionality, and may serve as a potential candidate therapeutic agent for tumors.

Chromatography, Affinity ; methods ; Escherichia coli ; genetics ; metabolism ; Humans ; Nickel ; Protein Folding ; Recombinant Fusion Proteins ; biosynthesis ; chemistry ; genetics ; isolation & purification ; Streptavidin ; biosynthesis ; genetics ; Tumor Necrosis Factor-alpha ; biosynthesis ; genetics