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Abstract:The immunoregulation effect to polysaccharides from Bifidobacterium spp. was investigated on the base of functional assessment standards of health food. Effects of the EPS on immunity were investigated by promoted the proliferation of spleen lymph cells, delayed type hypersensitivity reaction, the HC50 value and macrophage function assay in mice. Data showed that the EPS could obviously increase the ratio of swallowed chicken red blood cell by macrophage and the HC50 value in mice. However, no significant effect was found on the delayed type hypersensitive induced by sheep red blood cell, for only the low dose of 100 mg/(kg·d) EPS promoted the proliferation of spleen lymph cells. Bifidobacterium spp. EPS can certain immunomodulating function.

Abstract:In this study, we cloned genes coding the N-terminal region of PspA( pneumococcal surface protein A) from five of the most epidemic pneumoniae serotypes in China(FY01, FY05, FY6B, FY19F, FY23F). The obtained genes were respectively constructed into prokaryotic expression vector pET-27b(+), and then recombinant plasmids pET-pspA were respectively transformed into Escherichia coli BL21(DE3) for PspA expression. Induced with lactose, the five recombinant proteins were highly expressed and then purified by Ni-chelation chromatography. PCR with Family special primers was carried out to identify Family of PspA, and Clade of PspA was identified by means of bioinformatics according to the sequencing results. FY01rPspA, FY6B rPspA and FY19FrPspA belonged to Clade 1 of FamilyⅠ, FY05rPspA belonged to Clade 2 from FamilyⅠand FY23F rPspA belonged to Clade 3 from FamilyⅡ. Strong immunogenicity had been demonstrated in mice immuned with FY01rPspA at dose of 20 μg/mL each, of which antiserum antibody titer reached 1:210000. Result of western blotting showed that FY01rPspA antiserm reacted well with Y6B rPspAand FY19FrPspA but poorly with the other two, which suggested that cross-immunoreactivity was restricted in the same Clade. Mice immuned with FY01rPspA protected well against FY6B, while against FY05, FY23F was not effectively protected. The result in this study has much instructive significance for the development of a new efficient vaccine against pneumoniae.

Abstract:SYPS-062 was an L-serine producing strain stored by our lab, which could directly produce L-serine from sugar. According to morphology, physiological and biochemical identification and 16S rDNA sequence, SYPS-062 was identified as Corynebacterium glutamicum. And the effect of different carbon source for the SYPS-062 and Corynebacterium glutamicum ATCC13032 fermentation were studied. The experiment result showed that the sucrose was 60 g/L, the yield of L-serine and biomass reached the maximal value, 6.6 g/L and 8.1 g/L respectively.

Abstract:α-Hydroxynitrilelyase (HNL), a vital tool in the bio-synthesis of optically active cyanohydrin, which can catalyze carbonyls and HCN selective forming optically active cyanohydrin. HNL gene was amplified by PCR using pET30a-HNL as template and then ligated into pMD18-T vector. Validated HNL gene by sequenced was linked into pHMOXGαA vector. Positive recombinants grown in the medium containing 4 mg/mL of G418 were screened using PCR. Bio-active α-hydroxynitrilelyase expressed in Hansenula polymorpha after 0.5% methanol induction was obtained with 2.015 U determined by analysis of enzyme activity and SDS-PAGE.

Abstract:The glycerol fed-batch fermentations with a novel isolated 1,3-propanediol production strains, designed Klebsiella pneumoniae HR526 were performed in 5 L B. Braun fermenter. The result indicated that 91.47 g/L 1,3-propanediol was produced in 30 h. The extracellular metabolic flux was calculated, indicating PDO flux reached the maximum value in the mid logarithmic growth phase, whereas lactic acid flux reached the maximum value in the stable phase. This article also analyzed the key enzyme activity of PDO synthesis, the maximum PDOR activity was reached in the mid of the logarithmic growth phase, the ratio of the GDHt/PDO and GDHt/GDH is rather higher in the logarithmic growth phase than in the stable, leading to 3-HPA accumulation during the exponential growth phase. The production of PDO is closely related to cell growth.

Abstract:The glutamate dehydrogenase (EC.1.4.1.4) gene which amplified from the genome of Brevibacterium flavum GDK-9 by polymerase chain reaction was linked with pUCm-T for sequence alignment. Analysis of gdh sequences revealed that the whole sequence is 1927 bp, only one ORF existed, which used ATG as the initiation codon and coded a peptide of 448 amino acids with a calculated molecular weight of 48 kD. The comparability between the cloned gdh sequence to the reported sequence is high to 99.55%. Only the 1190th base mutation (C→A) lead to the change of amino acid sequence (Thr→Asn), the others are not. The recombinant plasmid pXG was then transformed into E. coli XL-Blue and Brevibacterium flavum GDK-9 which was induced by IPTG. SDS-PAGE analysis revealed that there was a clear induced protein band with molecular mass of 48.7 kD on expected position. Standard glutamate fermentations indicated that although the level of GDH increases the intracellular glutamate pool, the level of GDH has no influence on glutamate secretion.

Abstract:Lipase from Burkholderia cepacia complex is one of the most versatile biocatalyst and is used widely in many biotechnological application fields including detergent additives, the resolution of racemic compounds, etc. Based on the known whole genomic information of B. cepacia, both ampicillin and kanamycin were added to the TB-T media, the traditional selective media, to screen B. cepacia complex strains from rhizosphere soil samples. The single colonies on the plates with the modified TB-T media were then qualitatively determined the ability to produced the extracellular lipase in the rhodamine B-olive oil agar plates. Thirty-five strains of lipolytic pseudo-B. cepacia complex were isolated and the positive rate of lipolytic bacteria was 65%. Among them, 15 pseudo-B. cepacia complex strains with the tolerance to benzene, n-hexane and n-heptane at the concentration of 10% (V/V) were selected and identified by the recA gene sequence. All of the 15 lipolytic bacteria belonged to the B. cepacia complex.

Abstract:The temperature and salinity ranges for growth of twelve strains of marine Bdellovibrio-and-like organisms (BALOs) which affiliated with four different phylogenetic clusters in the family Bacteriovoracaceae and the lytic ability of them to six kinds of common shrimp pathogen vibrios were studied. The morphological characteristics of four representative strains were examined with transmission electron microscope. The results showed that the strains within the same cluster or sub-cluster had identical growth temperature or salinity. The growth temperature ranges of clusters IV, VI, IX and X strains were 20°C~35°C, 15°C~35°C, 15°C~40°C, 10°C~40°C, respectively, and the optimum growth temperature was about 30°C or 35°C. The growth salinity ranges of clusters IV, VI, IX and X strains were 5‰~40‰, 2.5‰~30‰, 5‰~60‰ and 5‰~60‰, and the optimum salinities were about 10‰, 5‰, 20‰ and 20‰, respectively. Three of the six kinds of vibrios tested were lysed by all the BALOs, while the others could only be lysed by some of the BALOs. However, differential lysis to some of the vibrios was also observed among BALOs within the same cluster. The four strains of different clusters all exhibited typical BALOs morphology, having vibrioid cell with a single polar flagellum.

Abstract:Toxic effects of L7 lyophilized powder of extracellular active components (L7-LPEAC), extracted from the Algae-lysing bacteria L7, on Chlorella pyrenoidosa were studied according to the changes of effective photosynthesis rate (EPR), the protein content, the chlorophyll a content and the MDA content of algae. The results showed that the growth of alga was promoted at low concentrations of L7-LPEAC (0.80 g/L, 1.25 g/L). The 96 h-EC50 and 120 h-EC50 upon Chlorella pyrenoidosa are 5.75 g/L and 2.55 g/L, respectively. The chlorophyll a content increased firstly and then decreased at high concentrations of L7-LPEAC (≥2 g/L), so did the protein content. Compared with the control group, there is a significant statistics difference (P<0.05) in the treatment groups (2.00 g/L, 3.13 g/L, 4.90 g/L), while extreme significant statistics difference (P<0.01) in the other treatment groups (7.67 g/L, 12.00 g/L) after 72 h. The inhibition rates of chlorophyll a content of Chlorella pyrenoidosa in all treatment groups reached over 60% after 120 h. For the remediation of eutrophic nature water body, it can not only kill the algae which caused the water eutrophication, but also maintain the balance of the aquatic ecosystem well by adding some proper concentrations of L7-LPEAC.

Abstract:Mechanisms of UV-B-induced apoptotic regulation in yeast Saccharomyces cerevisiae were studied. The results showed that UV-B irradiation indeed inhibited the growth of yeast cells as well as induced extensive apoptosis during 96 h experiment period. However, survival of 96 h irradiated cells remained 10% while most control cells finally dead after re-growth under UV-B irradiation for 12 d. And by exposed to 0.01 mol/L or 0.1 mol/L H2O2 for 30 min, survival rate of 24 h irradiated cells were 3.0-fold or 3.2-fold than control, respectively. By to heat shock for 30 min or 60 min, survival rate of 24 h irradiated cells were 3.5-fold or 9.0-fold than control, respectively.

Abstract:An effective RT-PCR method was developed to detect exogenous gene xylB transcript levels in Zymomonas mobilis CP4. Total RNAs without genomic DNA contamination were purified from wild-type and gene engineering strains, and were quantified to the same concentration. Then, cDNAs synthesis and PCR analysis of these samples were conducted by reverse transcription PCR. The optimal number of cycles was determined by observing amplification profile of target gene xylB and internal control gene 16s rRNA, and relative expression levels of xylB in various samples were analyzed by RT-PCR. The results indicated that the xylB transcript was not be detected in CP4, however that could be found in recombinant strains, in which xylB transcription abundance was similar. The enzyme assay furthermore confirmed that effective expression of the target gene. The method provided a useful and rapid tool for detecting transcript levels of target genes from various samples of Z. mobilis.

Abstract:The CBD-encoding region of family 7 cellobiohydrolases gene from the thermophilic fungus Chaetomium thermophilum was inserted into the upstream of the CelB gene from the hyperthermophilic bacterium Thermotoga maritime. The recombinant plasmid pHsh-CBD-CelB was expressed in Escherichia coli JM109. CBD-CelB fusion protein was purified by a simple heat treatment followed by DEAE-Sepharose FF column. The optimal temperature of CBD-CelB fusion protein for CMC activity was 90°C, and the fusion protein had the ability of binding crystalline cellulose. Enzyme activity of CMC and Avicel was increased.

Abstract:Soybean fungal root rot diseases are the most serious soybean disease problem in Heilongjiang province. The microbial seed dressing agent comprised of Bacillus strain B29 which showed strong antagonistic ability to the pathogens of Fusarium oxysporum, Pythium spp. and Rhizoctonia solani were applied to control soybean fungal root rot in field. The control efficiencies of the treatment T3, T4, T5, T6 on soybean fungal root rot after 35-day cultivation were 50.2%, 60.0%, 48.3%, 49.4% respectively, which were better than that of Carbendazim-Carbofuran-Thiram seed coating agent (T2). The second investigation after 50-day cultivation showed the control efficiency of T4 was the best (60.7%), while the control efficiency of T3 was not significantly different from T2. The control efficiencies of T5 and T6 were lower than that of T2. The microbial seed dressing agents were safe to soybean cultivation based on the investigation on seed germination rate, plant height, fresh weight and root nodules. The results based on 2-year demonstrating in field showed that soybean seeds germinating rate was more than 90% after seeds dressing by the microbial seed dressing agents, and the control efficiencies on soybean root rot diseases were 56.3% to 89.1%. It suggested that the microbial seed dressing agents were not only efficient to control soybean root rot diseases but also increased soybean productions.

Abstract:The effect of antibacterial active substances from Paenibacillus polymyxa Cp-S316 on the integrity of Escherichia coli K12 membrane was studied by determining the variation of OD260 values of bacterial suspensions. The results showed that the antibacterial substances could damage the Escherichia coli K12 membrane which leaded to the leakage of RNA, DNA and other cytoplasmic macromolecules. As an original strain, Paenibacillus polymyxa Cp-S316 was treated by ultraviolet mutagenesis. The mutated strain of A17 was obtained by prescreening of antibacterial active substances, screening and re-screening of shake flask culture. Compared with Cp-S316, the yield of antibacterial substances obtained from mutant A17 was increased by 91%. The subculture experiments indicated that the hereditary characteristic of high productivity of Paenibacillus polymyxa A17 was stable.

Abstract:In order to gain the strain which has high growth vigor and whose biomass concentration satisfy further fermentation to produce arachidonic acid, we took the yield of microbes olein and arachidonic acid as evaluative index, adopted twice ultraviolet mutation, determined the time of ultraviolet irradiation by single factor expriment, and then the content of the arachidonic acid was measured by GC. The results of this experiment showed that the power of viltalight lamp was 20 W, exposure distance was 30 cm and exposure time was 80 s, lethality of spores of Mortierella isabellina was 76.4%. After twice ultraviolet mutation and repeatedly screen, a mutation of Mortierella isabellina Z80s2-109 whose arachidonic acid concentration in biomass were 3.77 times of the control strains was obtained and genetic stability.

Abstract:An immunopotentiating compound has been isolated from the metabolites of Bacillus mycoides under the bioassay-guided isolation and identification for its immunopotentiating effect and chemical structure. The isolation and purification of the compound were consisted of macroporous adsorptive resins, silicagel chromatographic column and Sephadex G-200 chromatographic column. The immunopotentiating effect was assayed in every step isolation. At last, the only substance having the strongest immunopotentiating effect had been isolated and purified. Through the procedure consisiting of Ultra-Violet spectroscopy (UV), IR (Infrared Radiation), Time of Flight Mass Spectrum (TOF-MS), Nuclear Magnetic Resonance (NMR) and Element analysis, the possible structure of compound M had been identified as cyclic (Pro-Gly) dipeptide (C7H10O2N2) (Diketopiperazine). To be determined the immunopotentiating effect, the mice were treated by intraperitoneal injection of cyclic (Pro-Gly) dipeptide in the treatment group and distilled water in the control group. At the 14th day after the injection, the SOD activity and the phagocytosis activities reached the peak value and were significantly higher than those in control group. At the 21st day, the bactericidal activity reached peak value and was significantly higher than that in the control group. From the above results, we concluded that the main active component enhancing the immunity of mice was cyclic (Pro-Gly) dipeptide in the metabolism of Bacillus mycoides.

Abstract:We isolated 61 endophyte isolates from the bark of 4 plants, Ginkgo biloba L, Albizzia?julibrissin Durazz, Ailanthus sltissima (Mill) Swingle and Melia azedarach L. At the test concentration of 200 μg/mL, higher than 50% of antitumor activities were demonstrated with crude extracts from 45.9% of fungal culture in MTT assay. Six isolates, YX5, YX17, YX36, KL1, CC1 and CC5, still showed higher cytotoxicity at 50 μg/mL. No isolates from A.?julibrissin had inhibitory effect towards EC109 at the test concentration of 50 μg/mL; while about 15.8% of isolates from G. biloba were active. IC50 of the extract from the most active isolate YX5 against EC109, HONE1 and HeLa were 18.3 μg/mL, 3.6 μg/mL and 6.5 μg/mL, respectively. Our results indicate that endophytes from G. biloba could be regarded as a potent source of antitumor drugs.

Abstract:To study and optimize the fermentation parameters for expressing human-like collagenⅡduring E. coli high-density fermentation. The effects of pH, temperature, dissolved oxygen and induction instant on the cell growth and human-like collagenⅡproduction were investigated to optimize the fermentation conditions. The results demonstrated that the following conditions were beneficial for cell growth and foreign gene expression, controlling pH in phase induction at 6.8 and initial pH at 6.5, maintaining fermentation temperature and dissolved oxygen concentration was controlled at 34°C and 20% respectively, and implementing induction at the later logarithmic growth phase. Under the optimized condition, the cell density and human-like collagenⅡyield could reach 88.4 g/L and 14.2 g/L, respectively.

Abstract:To investigate the induction of apoptosis of mouse colonic adenocarcinoma CT26 cells by recombinant Clostridium difficile toxin B (rTcdB), CT26 cells were exposed to different concentrations of rTcd B. Inhibition of cell proliferation was measured by MTT assay. The activation of Caspase 3 was measured by colorimetric method. Cell morphological analysis and flow cytometry were performed to confirm cell apoptosis. rTcd B inhibited the proliferation of CT26 cells in a time- and dose-dependent manner. Caspase 3 activity in CT26 cells was elevated remarkably after rTcd B exposure for 6 h, 12 h, 18 h or 24 h, as compared with the control group. Morphological changes were observed by fluorescence microscopy. The exposure of rTcd B to CT26 cells induced a time- and dose-dependent apoptotic cell death as determined by flow cytometry analysis. The results showed that recombinant Clostridium difficile toxin B induced apoptosis of CT26 cells.

Abstract:An actinomycete strain P-13, with antimicrobial activity against muskmelon bacterial spot pathogens, was isolated from the muskmelon rhizosphere soil samples in Xinjiang. The strain P-13 was identified as Streptomyces rochei based on morphological, physiological characteristics and 16S rRNA sequence analysis. The agar diffusion bioassay showed that the diameter of inhibition zone against Acidovorax avenae subsp. citrull BFB and Pseudomonas syringae pv. Lachrymans P4 was above 19 mm and 17 mm, respectively. The antimicrobial substances obtained from strain P-13 were demonstrated to be alkaline and water-soluble compounds according to paper chromatogram analysis and exocellular metabolites. Furthermore, it was stable to be treated by 100°C for 10 min, pH 6 for 6 h, or ultraviolet treatment for 7 h. Moreover, it was insoluble in organic solvents, such as petroleum benzine, diethyl ether, and acetic ether.

Abstract:The development of odor-control technology currently used in domestic and overseas was reviewed in this paper, various technique used to control the odor gases generated from municipal wastewater treatment plants were introduced, and the advantages and disadvantages of each present method of removal technology were compared. The compared result indicated that water washing and spewing was easy to do but had low removal rate. The method of chemical absorbing and active carbon can remove most of the odors, but the expense is also very high. Biotechnology was economical and effective. It can keep high removal rate with no secondary contamination and will be extensive applied in the ordor treatment of municipal wastewater treatment plants in future.

Abstract:The rhizosphere is the narrow region of soil that is directly influenced by roots and associated soil microorganisms. Research on rhizosphere microbe is essential for microbial ecology and industrial biotechnology. However, the inability of culturing most rhizosphere microorganisms (around 99%) in the laboratory obviates the research progress. In recent years, there is enormous advance in applying non-culturing techniques based on molecular biology and genomics to the study of rhizosphere microbial diversity and plant-microbe interactions. This review discusses recent findings and future challenges in the study of rhizosphere microbe, with brief comment of our Taxus rhizosphere study and various novel techniques.

Abstract:Commercial production of bioethanol from lignocellulosic hydrolysates requires efficient fermenting strains. The abilities of the strain to converting all types of sugars in the hydrolysate to ethanol in high yield and to effectively tolerating/metabolizing inhibitors are necessary. Genome shuffling is a novel method for breeding, and it has been applied in pharmaceutical and food industry. This review summarized the technique of genome shuffling including principle, process, applications and its prospect for strains improvement in ethanol production from lignocellulosic hydrolysates.

Abstract:mazEF, which is a toxin-antitoxin system located in bacterial chromosome, can mediate programmed cell death induced by various stressful conditions. In this review, the genetic structure, physiological and biochemical function of mazEF system were outlined, cellular signals and the regulation of cell factors involved in the process of cell death were introduced, and the controversy on the theory of mazEF-mediated programmed cell death was discussed. Based on the present results, It is pointed out that some questions about bacterial programmed cell death should be taken into more consideration.

Abstract:According to teaching practice of “microbial engineering”, teaching reforms, such as conformity and optimization of the curriculum system, improvement of teaching methods and the construction of multilevels experimental teaching system, are investigated in this paper, in order to improve the teaching quality and enhance the overall quality and the abilities of operation and innovation of students.

Abstract:In order to cultivate the students’ abilities of thinking and practicing and enhance their comprehensive abilities in microbiology experiment, the authors try to search for some new teaching ways and assessment methods in microbiology experimental teaching and also attempt to make some improvement in the textbook and adjustment in teaching schedule so as to develop the students into specialized talents.

Abstract:Thirty strains of seven Candida species from CICC(China Center of Industrial Culture Collection)were studied. The strains were differentiated by ITS1 region PCR-SSCP fingerprinting analysis and ITS2 region PCR-SSCP fingerprinting analysis. Results showed that both ITS1 region and ITS2 region were able to differentiate the seven species of Candida clearly. Contrasting the maps and effects on the identification of Candida species of ITS1 region with that of ITS2 region, result indicated that on the identification of Candida species the application of ITS2 region was better than ITS1 region.

Abstract:A rapid LAMP detection method with primers designed on genus-specific region identified in the gyrA gene was established in this assay. All four Campylobacter jejuni from different sources were detected positive and fourteen non Campylobacter bacteria were negative, which shows excellent specificity of the primers. Compared with plate count and PCR method, the LAMP method and the PCR method had equal sensitivity, which were three orders of magnitude higher than plate count. In this assay, we also found out that the treatment of DNase could reduce the dead bacteria DNA effectively. The LAMP detection on chicken indicated relatively good result on detection of Campylobacter jejuni combining with treatment of DNase.

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