Abstract:[Objective] We studied physiological, biochemical properties and metabolites of Thermoanaerobacter mathranii SC-2 from oil-field water in Shengli oilfield. [Methods] Strain SC-2 was isolated by Hungate anaerobic technique. Through physiological, biochemical and phylogenetic analysis, the strain was identified. Metabolites were analyzed by gas chromatogram. [Results] The cells were Gram-negative, rod-shaped, spore-forming. Growth was observed in the temperature range from 40 to 75℃(optimum 70℃) and pH range from 5.5 to 9.5 (optimum 6.5). The isolate grew in the presence of 0%~5% NaCl with an optimum without NaCl at pH 7.0 and 65℃. Strain SC-2 used many carbohydrates as carbon sources, including glucose and xylose. Metabolites of glucose were ethanol, acetate, propionate, lactate, CO2 and H2. Based on 16S rDNA studies, strain SC-2 was most close to T. mathranii subsp. mathranii11246T with 99.85% similarity. More ethanol and acetate were produced at initial pH 8.0 than yields at other pH. Yeast extract could significantly increase ethanol and acetate yields. In addition, ethanol (4%) added in the medium obviously inhibited its growth. [Conclusion] Strain SC-2 was extremely thermophilic, halotolerant anaerobe.

Abstract:[Objective] To know halophilic actinomycetes and their enzyme-producing ability from mud volcano in Usu County, Xinjiang, China. [Methods] Soil samples in mud volcano were isolated with five isolation media containing 5% and 10% NaCl (w/v) with dilution-plate method. The activities of lipase, galactosidase, amylase, esterase and cellulase from isolated strains were qualitatively detected by using five selective media. Base on morphological characteristics, test of salt tolerance, screening of enzymatic characters and sequencing of 16S rDNA gene, strains were selected for phylogenetic analysis. [Results] A total of 43 halophilic actinomycetes and 3 extreme halophilic actinomycetes were obtained. Screening results for enzyme activity showed that 4 halophilic actinomycetes produced lipase, 30 strains produced galac-tosidase, 27 strains produced amylase, 6 strains produced esterase, 4 strains produced cellulose and 1 strain produced 4 enzymes simultaneously. According to 16S rDNA sequence analysis, 24 of 25 detected sequences were affiliated with Nocardiopsis, and the other one was Streptomyces. In the phylogenetic tree, strain 10006 producing 2 enzymes and No-cardiopsis exhalans (AY03600)showed 96.64% similarity (less than 97%), which indicated that strain 10006 was a possi-ble new species. [Conclusion] There are plenty of galactosidase or amylase-producing halophilic actinomycetes and higher enzyme-producing diversity of Nocardiopsis in Usu mud volcano. In addition, there are potential microbial re-sources in this extreme environment.

Abstract:[Objective] We tested the antimicrobial susceptibility of 390 Salmonella isolates. We also studied the relationship between plasmids in some multidrug resistant Salmonella isolates and the antibiotic resistance profile of their hosts, as well as conjugation test of some multidrug resistant Salmonellas. [Methods] Salmonella strains were isolated by using selective cultures, putative Salmonella was confirmed by PCR. Antimicrobial susceptibility was tested according to the National Committee for Clinical Laboratory Standards. Plasmid of some representative multidrug resistant strains was isolated by using QIAGEN Plasmid Mini Kit and digested with HindⅢ. The plasmid profiles were acquired by gel electrophoresis and analyzed by DPS. The conjugation test was done to illustrate the function of plasmid during the antibiotic resistance transfer. [Results] Of the Salmonella isolates, 58.2% were resistant to tetracycline, followed by resistance to streptomycin (42.8%), kanamycin (39%), ampicillin (38.2%), cefoxitin (27.2%), chloramphenicol (26.9%), gentamicin (21%), ceftriaxone (19%), amoxicillin-clavulanic (18.2%), trimethoprim-sulfamethoxazole (17.9%), ceftiofur (14.6%) and nalidixic acid (12.3%). There was no strict corresponding relationship between antibiotic resistance profile of the host Salmonella and its plasmid profile. The conjugation frequency of the plasmid was from 2.4×10-4 to 5.6×10-1. [Conclusion] Antimicrobial resistance is common in foodborne Salmonella, direct relativity does not exist between the homology of plasmids and their hosts’ antibiotic resistance phenotype, antibiotic resistant genes in the plasmid can transfer from donor to the recipient in interspecies and intraspecies with high frequency accompanying conjugation.

Abstract:[Objective] We cloned the promoter of glycerol-3-phosphate dehydrogenase gene(CgGPD) from the Candida glycerinogenes, and studied its functional regulation under high osmotic stress condition. [Methods] We amplified the 950 bp promoter of CgGPD from C. glycerinogenes and the green fluorescent protein gene (gfp) from pCAMBIA1302 vector by PCR and introduced them into a modified vector pYX212-zeocin simultaneously. The recombinant plasmid pYX212-zeocin harboring both the promoter of CgGPD and gene gfp was transformed into S.cerevisiae W303-1A by electroporation. In the medium containing glucose with different concentrations for culturing the recombinant strain S. cerevisiae W303-1A--GFP the green fluorescence was detected by fluorescent microscopy. [Results] The gene gfp was functionally expressed under the control of the promoter of CgGPD in S. cerevisiae. Furthermore, the expression of the gene gfp at different level was conducted by the different osmotic stress for the recombinant strain. The green fluorescence was less intensive when the concentration of glucose was low for culturing the recombinant strain, but it became much more intensive when the concentration of glucose increased. [Conclusion] The promoter of CgGPD is an inducible promoter that can be induced significantly by the high concentration of glucose. The promoter will facilitate further studies on the mechanism of glycerol synthesis from C. glycerinogenes WL2002-5 under osmotic stress conditions.

Abstract:[Objective] To identify genes involved in multidrug resistance in Pseudomonas aeruginosa. [Methods] We constructed and screened a transposon mutation library and obtained mutants that exhibited decreased drug resistance. The insertion sites of the transposon were identified by arbitrary PCR and subsequent DNA sequencing. [Results] Two mutants which became more susceptible to several antibacterial agents than the wild type were identified as mutants with transposon insertion in gene PA2580 and PA2800 respectively. The functions of these two genes are described as unknown in P. aeruginosa database. [Conclusion] PA2580 and PA2800 are involved in drug resistance, possibly through their roles in redox reactions and cell envelope biogenesis.

Abstract:[Objective] We developed recombinant fowlpox viruses (rFPV) coexpressing chicken IL-18 and H5 AIV HA. [Methods] Recombinant expression plasmid pSYHA/IL-18 was constructed by cloning chicken IL-18 into transfer plasmid containing HA gene and transfected by lipofectamine on the chicken embryo fibroblasts cell (CEF) pre-infected with S-FPV-017. By selecting blue plaques on the CEF overlaid with agar containing X-gal, recombinants fowlpox virus rFPV-HA-IL-18 were obtained, and identified by PCR. [Results]The recombinant fowlpox viruses contained chicken IL-18 and HA gene and had stable genetic properties. The expression of HA was detected in the recombinant virus-infected CEF by indirect immunofluorescence using antibody against AIV. The expression of chicken IL-18 was detected by MTT in the recombinant virus-infected CEF fluid. The chickens vaccinated with recombinant fowlpox virus rFPV-HA-IL-18 and rFPV-HA had detectable hemagglutination inhibition (HI) antibody at 7 days post-vaccination, and HI antibody titers rose to peak at 14 days post-vaccination. No HI antibody was detected in the control or fowlpox virus immunized chickens before or after immunization. The chickens vaccinated with rFPV-HA-IL-18 had higher HI antibody titers than the chickens vaccinated with rFPV-HA. [Conclusion] Development of recombinant fowlpox virus (rFPV- HA-IL-18) had strong biological activity.

Abstract:[Objective] Prolonged infection with Hepatitis B virus (HBV) has been recognized as a major factor for hepatocellular carcinoma (HCC). In this study, we studied the host protein Mre11 fluctuations after HBV infection which might be eventually contributed to cell transformation. [Methods] Western Blot was monitored to detect the expression of Mre11, and RNA interference was used to downregulate protein expression. Then, Ligation mediated PCR was used to detect the level of DNA double strand breaks. [Results] Mre11 protein was downregulated when HBV infection occured, and the downregulated expression was also seen in HCC tissue. By RNA interference, we found that Mre11 knockdown caused DNA instability. [Conclusion] Mre11 expression downregulation contributed at least partially to cell transformation caused by HBV infection.

Abstract:[Objective] During a screening for free radical scavengers from metabolites of entomogenous fungi, we fond a fermentation broth of the strain RCEF 0881 of Hirsutella sp. exhibited strong radical scavenging activity. To make clear of the constituents of the active compounds, and prepare some pure active compound for further structure identification we launched this study. [Methods] We used organic solvent for active compounds extraction. DPPH-TLC and DPPH- Mi-croplate assay were used for activity analysis. Components analysis was carried out on a HPLC-DAD-HRMS, and bioac-tive compound preparation on a preparative RP-HPLC. [Results] Our extraction tests showed that ethylacetate was the best solvent for the bioactive constituents extracting. HPLC-DAD-HRMS-DPPH assay revealed that the molecular for-mular of the radical scavengers existed in the extract were possibly C7H6O4, C8H8O3 and C12H14N2O. From the chroma-tographic and Uv properties, and the MS fragments, and database consulting, the compounds could be deduced as dihy-droxybenzoic acid, methyl-hydroxyl benzoic acid, and an alkaloid, however, the structures are still needed to be confirmed. The pick area of HPLC and MS showed that the compound C12H14N2O was the main component of the extract. It was iso-lated via activity directed fractionation. The activity of the prepared compound was confirmed with DPPH-TLC assay and its purity was confirmed with HPLC-DAD-HR-ESIMS. The occurance of the three active compounds in entomogenous fungi was revealed for the first time.

Abstract:[Objective] To study the mechanism of the differences in cell growth and acetate production between Escherichia coli DH5a and its acetate-tolerant mutant DA19. [Methods] Based on our previous analysis of the proteome and mass and energy balances for both strains, we first amplified the genes of atpIBEFHAGDC, purHD, and ndh, their regulation regions, as well as the regulation region of nuoA-N in both strains by PCR, and compared the sequences. Then, we constructed the recombinant strains DH5a (pTrc99a-atpA), DH5a (pTrc99a-purHD)?and DH5α (pTrc99a-ndh) to study the effect of overexpression of atpA, purHD and ndh in DH5a on growth. [Results] The sequences of the structural genes and the regulation regions in the two strains were completely identical with those of E. coli K-12. Overexpression of atpA, purHD and ndh separately in DH5a improved cell growth to some extent. [Conclusion] Overexpression of ndh, purHD and atpA resulted in improved growth of DH5a, but the growth of these recombinant strains was still lower than that of DA19, indicating the importance of global regulation to the cellular metabolism.

Abstract:[objective] In order to obtain high yield mutant strains for the industrial bioconversion of succinic acid, we analyzed the metabolic networks of the strain Actinobacillus succinogenes S.JST in the course of screening and breeding. [methods] We previously identified the wild-type strain by API biochemical reactions and 16S r RNA sequence analysis. Following the discussion of the metabolic pathway, we calculated the flux by matrix and disturbed the node by intermediate. [results] A succinic-acid-producing strain S.JST isolated from bovine rumen was identified as Actinobacillus succinogenes. Enzyme determination showed that the activities of phosphoenolpyruvate carboxykinase and malate dehydrogenase were very high. Metabolic flux from parent strain indicated that the flux of by-product ethanol was 1.51 mmol·g-1·h-1 in the second place of those end products. After being mutated, the alcohol dehydrogenase activity of the mutant-strain S.JSTA decreased markedly, furthermore the flux of succinic acid increased by 34% and the flux of ethanol decreased by 93%. By analyzing the Adh gene, we found a mutated site. Bioinformatics showed that the corresponding amino acid sequence acted as the key active site binding with NADH. [conclusion] In succinic acid synthesis, directed breeding method was effective for improving the whole cell metabolism of Actinobacillus succinogenes, and succinic acid yield was increased.

Abstract:[Objective] The mechanism of L-threonine biosynthesis and its impact factors were explored by metabolic flux analysis. [Methods] The metabolic flux balance model of L-threonine synthesis by Escherichia coli was established. Based on this model, the practical and optimal metabolic flux distribution in the middle and late period under different dissolved oxygen concentrations were determined with the linear program planted in MATLAB software. [Results] Data indicated that 25.5% of carbon sources were consumed by HMP pathway, resulting in a conversion rate of 33.9% to L-threonine with a 5% dissolved oxygen concentration. With dissolved oxygen concentration of 20%, 58.08% carbon resources entered HMP pathway, giving rise to a 46.5% conversion rate. [Conclusion] Compared to the optimal metabolic flux with a carbon conversion rate of 88.23%, glucose-6-phosphate isomerase should be activated by genetic manipulation and fermentation control in order to elevate the HMP pathway flux, and flux towards aspartate amino acids family could be enhanced by increasing phosphoenolpyruvate carboxylase reaction rate. These may lead to a decrease in TCA flux and byproducts, and consequently the L-threonine biosynthesis would be promoted.

Abstract:[Objective] The aim of this manuscript was to illuminate the effect of NADH oxidation pathway on the glycolytic rate and the pyruvate productivity. [Methods] The noxE gene encoding a water-forming NADH oxidase from Lactococcus lactis, was expressed in a pyruvate producing Torulopsis glabrata CCTCC M202019. A mutant strain T. glabrata-PDnoxE, with specific NADH oxidase activity of 34.8 U/mg protein, was obtained. [Results] During batch fermentation with 100 g/L glucose in the medium, the dry cell weight, the glucose consumption rate and pyruvate production rate were 168%, 44.9% and 12% higher than that of the parent strain, respectively. Only 2.5 g/L residual glucose was detected in the fermentation broth after 36 h culture, then 50 g/L glucose was supplemented to the culture broth and the concentration of pyruvate increased to 67.2 g/L. As the result of NADH oxidase overexpression, the intracellular NADH, NAD+ and ATP concentrations of the mutant and the parent strain were determined, the NADH and ATP content decreased 18.1% and 15.8% respectively, while the NAD+ concentration increased 11.1%. [Conclusion] The increasing of intracellular NAD+ concentration can efficiently enhance the rate of glucose consumption and the pyruvate production.

Abstract:[Objective] To enhance the production and bioactivity of recombinant human disintegrin domain of ADAM15 (A Disintegrin and Metalloproteinase 15) (rhADAM15) in Escherichia coli. [Methods] The expression host was chose and the recombinant expression plasmid pGEX-ADAM15 was constructed, based on analysis of the cDNA sequence of rhADAM15. [Results] (1) 298 mg/L GST (Glutathione-S-transferase)-ADAM15 and 42 mg/L rhADAM15 were achieved when choosing E. coli Rosetta (DE3) as the expression host that could supply additional tRNA for rare codons. (2) The GST-ADAM15 expression level increased to 326 mg/L after changing the rare codon GGA (Gly425) to GGC by PCR (Polymerase Chain Reaction)-based site-directed mutagenesis. (3) The rhADAM15 concentration increased to 57 mg/L by deleting the “Pro-Glu-Phe” at the GST-ADAM15 junction to improve the thrombin cleavage efficiency. (4) Finally, by combinational introduction of the favorable codons and suitably eliminating of certain amino acid sequence, rhADAM15 concentration reached the highest level (68 mg/L). [Conclusion] The high expression of heterologous protein could be achieved by releasing rare codon usage and amino acids residues restriction.

Abstract:[Objective] To isolate and identify a novel strain with cis-epoxysuccinate hydrolase (CESH) activity and to optimize its enzyme production. [Methods] The isolated strain was identified by electron microscopy, Biolog gram negative (GN) test, G+C (guanine plus cytosine) content measurement and 16S rDNA sequence. The purified enzymatic biotransformed product was identified by IR, 1H-NMR, 13C-NMR, MS and optical rotation analysis. Then the fermentation conditions for CESH production were optimized. [Results] A novel CESH-producing strain was isolated for biotransforming cis-epoxysuccinate to D(-)-tartaric acid. It was assigned to genus Bordetella and named Bordetella sp. BK-52. The optimal conditions were found to be 30°C, pH 7.0, fermentation time 36 h, car-bon source of saccharose, inorganic nitrogen source of ammonium sulfate and enzyme inducer of disodium cis-epoxysuccinate. Under these conditions, the maximum CESH activity reached 764 U/g biomass. [Conclusion] The isolated Bordetella sp. BK-52 provided a new alternative for biosynthesis of D(-)-tartaric acid from cis-epoxysuccinate.

Abstract:[Objective] Examining bacterial diversity in an oil reservoir of Shengli oil field by both culture-independent molecular technique and enrichment method. [Methods] The heterotrophic bacteria, hydrocarbon-degrading bacteria and sulfate-reducing bacteria were enriched from S12-4 oil-well samples by the corresponding media. Then the genomic DNAs of the enrichments were extracted and the 16S rRNA gene clone libraries were constructed. [Results] The phylogenetic analysis revealed that the bacterial 16S rRNA gene clone libraries of the 3 enrichments were dominated by clones of Thermotoga, Thermaerobacter and Thermotoga, respectively. Sequences of the other co-dominant clones observed only in the enrichments of hydrocarbon-degrading bacteria and sulfate-reducing bacteria were, respectively, associated with Marinobacter and Moorella. The uncultured 16S rRNA gene library was also generated directly from total DNA of S12-4 oil-well samples by bacterial primer set. Sequence analysis of this bacterial library indicated that a large percentage of clones were highly related to the genus Pseudomonas and the dominant species emerging in the enrichment samples had a very low content in the tested oil reservoir. The significant difference of the bacterial composition between the samples obtained from independent-culture method and enrichment method implies that the specialized nutrient may lead to a distinctive selection of dominant organisms. [Conclusion] Through culture-dependent and culture-independent methods, we acquired important information on the bacterial diversity of ShengLi oil reservoir. These results may expand our understanding of the microbial diversity of oil reservoir and provide useful information for MEOR(microbial enhancement of oil recovery).

Abstract:[Objective] To study the characteristics of the Nitroso-bacteria in one-step completely autotrophic nitrogen removal process. [Methods] We took samples from activated sludge of the one-step completely autotrophic nitrogen removal reactor. Through four times of enrichments and isolations, one Nitroso-bacteria named N1 was obtained. We identified this strain by microscope and 16S rDNA analysis and studied the effect of pH, temperature as well as ammonium concentration on the metabolism of N1. [Results] Of the N1 nucleotides, 97%, 96% and 96% were identical with the conserved fragment of Nitrosomonas sp. NL7 (AY958677), Nitrosomonas AS1 (EF016119) and Nitrosomonas sp. Is32 (AJ621027) respectively. The optimal temperature and pH of N1 were 8.0 and 30℃ and sufficient dissolved oxygen was demanded. In addition, no restraint effect was turned up in the ammonium concentration of 80-800 mg/L. [Conclusion] N1 was identified as Nitrosomonas sp. Compared with the relative references, this strain was ammonium-tolerant.

Abstract:[Objective] To study the effect of troglitazone on autophagy and programmed cell death of HeLa cells. [Methods] Fluorescence microscopy, electron microscopy, western-blotting and flow cytometry were used. [Results] Troglitazone increased the autophagosome formation of HeLa cell as observed by fluorescence and electron microscopy. Immunoblotting further confirmed cell morphology observation upon the reagent stimulating by detecting an enhanced expression of LC3, an autophagic related gene. Besides apoptosis, troglitazone triggered a necrotic cell death in flow cy-tometry analysis. [Conclusion] Troglitazone induced mixed type of cell death in HeLa cells.

Abstract:[Objective] We clarified the genetic structure and phylogenetic relationship of the aroA gene in Haemophilus parasuis, a gene encoding 5'-enolpyruvylshikimate-3-phosphate synthetase. [Methods] We used PCR and bacterial genome walking techniques to determine the aroA gene locus in the genome of H. parasuis serovar 5. Then we sequenced the aroA gene of H. parasuis reference strains and local isolates. All aroA gene sequences from H. parasuis, other members of the family Pasteurellaceae, Escherichia coli and Salmonella typhimurium were analyzed for phylogenetic relationship. [Results] An approximately 3.7 kb fragment containing the whole aroA gene was obtained from the genomic DNA of H. parasuis serovar 5. Sequence analysis showed that the size of the whole aroA gene was 1314 bp, encoding a 437 aa product with molecular weight 47.9 kDa. The sequence immediately upstream of the aroA gene shared high similarity with phosphoenolpyruvate carboxylase gene. The whole aroA gene fragment could be amplified from all strains of H. parasuis serovars with a size of 1476 bp. The nucleotide sequence similarity was above 97.7% in H. parasuis serovars. The simi-larity was 70.6%-78.9% between H. parasuis serovar 5 and other members of Pasteurellaceae. The similarity between H. parasuis serovar 5 and E. coli or S. typhimurium was 66.4% and 67.2% respectively. [Conclusion] No obvious difference was found in the aroA gene sequence of H. parasuis reference strains (15 serovars) and 3 local isolates. The nucleotide sequence of the aroA gene was well-conserved within Gram-negative organisms.

Abstract:[Objective] To identify putative lipoproteins of Streptococcus suis serotype 2 (SS2). [Methods] First, I used the ScanProsite feature of Prosite and the website Database Of bacterial LipOProteins(DOLOP) to identify putative lipopro-teins in the recently published genome of SS2 strains 05ZYH33, then validated the identified putative lipoproteins to ex-clude the false positive by a ‘majority vote’ approach following analysis with standard tools for signal peptide verification. Finally, putative functions were attributed to individual lipoproteins by reference to the identification of conserved do-mains of InterPro. Homologous proteins were identified by unfiltered BlastP homology searches (including conserved domain detection). [Results] Among the 38 identified putative lipoproteins, 34 were validated as lipoprotein and 4 as false positive lipoproteins. The largest functional category (16/34, 44%) of lipoproteins was that predicted to comprise of sub-strate binding proteins of ATP-binding cassette transport systems. Other roles included lipoproteins that participated in adhesion (YP_001197698 and YP_001198710), protein export and folding. [Conclusion] Lipoproteins contributed to the physiology of SS2 and influenced its virulence.

Abstract:[objective] We developed an indirect enzyme-linked immunosorbent assay(ELISA) by the recombinant VP1protein of duck hepatitis virus (DHV) expressed in Escherichia coli to detect antibodies against DHV. [Methods] The VP1 gene of DHV was amplified by reverse transcription-polymerase chain reaction (RT-PCR) and cloned into pMD18-T and pET-32a vectors to get a prokaryotic expression vector pET-32a -VP1. DHV VP1 gene was expressed and analyzed. A method of enzyme-linked immunosorbent assay (ELISA) was developed and studied. [Results] We obtained the recombinant plasmid pET-32a-VP1 with correct sequence and orientation. DHV VP1 gene was expressed in E. coli BL21(DE3) at a high level and had good immunoreactivity by SDS-PAGE and western blot. The optimum working concentration of antigen was 5.0 mg in 100 mL per well, the working concentration of serum samples was 1∶100 dilution and the critical value was OD450≥0.302 for the ELISA assay. The rate of coincidence of ELISA and virus neutralization test(VN) was 97.5% by detecting 80 serum samples. The method was specific, sensitive and could be applied for antibody detection of maternal antibody and the rule of antibody growth and decline in immunizing ducklings. [Conclusion] The ELISA method developed by the purified recombinant protein could be used to detect antibodies against DHV.

Abstract:[Objective] The aim of the study is to establish in vitro cell line with stable and effective 3Dpol gene expression, so as to study the biological function of foot-and-mouth disease virus (FMDV ) 3Dpol and foot-and-mouth disease (FMD) gene engineering vaccine. [Methods]FMDV 3D gene was amplified from pMD18-T-3D and inserted into pGEM-Teasy vector. By NotI/BamHI digestion, 3D gene with NotⅠ/BamHⅠsite was inserted into NotⅠ/BamHⅠcloning site of the pBPSTR1 retroviral vector in order to obtain recombinant retroviral vector pBPSTR1-3D. The artifical retroviral viruses were obtained by both pBPSTR1-3D and pVSV-G envelope vector into the Gp2-293 package cells using Lipofectamine 2000. The BHK-21 cells were infected by artificial retroviral particles with 8 mg/mL Polybrene. The positive cell clones which genomes contained the 3Dpol gene were continually selected using puromycine and was regulated by tetracycline for 12 days .The single clone highly effective expressing 3D fusion protein was obtained by seeding the cells into 96-well plates with one cell per well. [Results] By using retroviral gene transfer technology, the 3Dpol gene was integrated into the chromosome of BHK-21cells, then under selection pressure, the cell lines stably expressing 3D were established. Finally, a cell line stably expressing the 3D fusion protein was established. The fusion protein was confirmed to be expressed correctly by Western-blot. The transfected genes in the cell line were consistently expressed during 35 passages of the host cells. [Conclusion] Transgene cell strain stably carrying exogenous gene in subsequent passaging was successfully constructed. It provide a good experimental tool for the biological function of FMDV 3Dpol and FMD gene engineering vaccine research.

Abstract:[Objective] We studied biologic characteristics of Stx2-converting phage induced from Escherichia coli O157 by mitomycin C. [Methods] Eight Stx2-converting phages were isolated from E. coli O157 and identified by PCR. The phage particles were purified and phage DNA was extracted. Random priming digoxin (DIG)-labeled stx2-specific gene probe was prepared for Southern blot. The morphology of these phages were studied by electron microscopy. Protein profiles were analyzed by Sodium Dodecyl Sulphate PolyAcrylamide Gel Electrophoresis (SDS-PAGE). Restriction fragment length polymorphism (RFLP) was used to confirm the size, type, and polymorphism of the purified phage genome. [Results] These 8 phages were confirmed Stx2-converting phage and DIG-labeled probe was highly specific. All phages had a regular hexagonal head and a short tail, belonging to Podoviridae phage families. The Stx2 phages had genome sizes in the range of 48 to 65.3kb, consisting of dou-ble-stranded DNA. The restriction fragment length polymorphism of these phages showed different groups, although the SDS-PAGE protein profiles of these phages were similar. [Conclusion] These 8 Stx2-converting phages with similar morphol-ogy belonged to Podoviridae phage families. The protein profiles were highly identical. We could differentiate these Stx2-converting phages according to their restriction fragment length polymorphism patterns.

Abstract:Glucose is degraded to pyruvate via the so called “central metabolic pathways” that play vital roles in the carbohydrate and energy metabolism of organisms. Some variances to the classical glycolytic pathways in bacteria and eukarya are presented in the glycolysis of archaea. Results from biochemical, genomic and metabolomic studies indicate that some novel and characteristic enzymes are involved in the archaeal Embden-Meyerhof (EM) and Entner-Doudoroff (ED) glycolysis pathway. The ED pathway in archaea is divided into two sub-routes-the semi-phosphorylative and non- hos-phorylative Entner-Doudoroff pathways. The unique glycolysis pathway in archaea is different from those in bacteria and eukarya in metabolic route, enzyme, regulation site, and energy transformation. These characteristics show the ability of these extremophiles to evolve flexible metabolic pathways in the extreme life environment. We reviewed recent advances in the ED glycolytic pathway of archaeon concerning enzymes, regulation and energy transformation. The potentials of glycolysis pathway in archaea were also discussed.

Abstract:The microbial secondary metabolites are always the main source of the natural drugs. The historical paradigm of the deep ocean as a biological ‘desert’ has shifted to one of a ‘rainforest’ owing to the isolation of many novel microbes and their associated bioactive compounds. A high quality microbial and its natural product library are crucial for successful drug and other screenings. However, how to build up the library efficiently is still faced with many bottlenecks. To overcome the difficulties and limitations, we reviewed the following strategies: (1) diversifying microbial sources and dereplication; (2) diversifying gene sources and dereplication; (3) diversifying microbial metabolite sources and dereplication; (4) novel methods and technologies for bioactive secondary metabolites, especially the high-throughput synergy screening for multi-target drugs. Bioactive compounds isolated using the above chemical microbiology strategies have not only shown importance in biotechnological and pharmaceutical applications but also increased our understanding of the diversity of microbe, ecosystem functions and the exploitable biology.