Abstract:In this work，the enhancement of unfolded protein response transcription pathway at different cultivation temperatures were integrated as a novel protein secretion strategy to improve extracellular enzyme concentration and specific productivity of heterologous glucose oxidase （GOD） in Pichia pastoris. Efficient production of heterologous proteins in P. pastoris was often limited by metabolic and intrinsic stresses such as folding rate and ER stress by accumulation of unfolded protein. The results showed that enzymatic activity of engineered secretion strain PP-G-HAC1，co-expression of UPR activating transcription factor Hac1p from P. pastoris，was up 161 U/mL in shake flasks， 34% higher compared to control strain. Furthermore，the effects of different cultivation temperatures （22、28 ℃） were also investigated. The highest GOD activity and extracellular concentration at 28 ℃ in PP-G-HAC1 were 1，008 U/mL and 14.43 g/L，respectively，3.12-fold increase than that of control strain.

Abstract:L. plantarum FS5-5，isolated from traditional home-made fermented soybean in northeast of China，has been considered as a new salt tolerance bacterium. Proteomics research were carried out to identify proteins expression of L. plantarum FS5-5 grown at different NaCl concentrations of 0% and 3% in this study. Cytosolic proteins of mid-exponential phase were resolved and further analyzed by two-dimensional gel electrophoresis. The results showed that sixteen protein spots were found with statistically significant differences.Twelve of total protein spots were identified as UDP-N-acetylglucosamine diphosphorylase，glucose-6-phosphate 1-dehydrogenase，UDP-N- acetylmuramoylalanine-D-glutamate ligase，adenylosuccinate synthetase，cysteine aminopeptidase，prophage protein，pyruvate kinase，ABC transporter（iron-sulfur cluster assembly protein），bile salt hydrolase，glutamine synthetase，6-phosphogluconate dehydrogenase，Xaa-Pro dipeptidase using matrix-assisted laser desorption /ionization timeof-flight mass spectrometry （MALDI-TOF/MS），indicating the complex response of lactic acid bacteria to salt stress．

Abstract:To promote the application of α-1, 6-mannose transferase (OCH1p) and α-1, 3-mannose transferase (ALG3p) double mutant strain of Pichia pastoris X33 (och1 alg3 mX33) as host for protein expression and to lay the foundation for the further glycosylation of the double mutant strain, this study wants to increase the growth rate as well as the ability to adapt of the double mutant strain through ultraviolet mutagenesis and Atmospheric and Room Temperature Plasma (ARTP) mutagenesis methods. At last, after four rounds of screening, a double mutant strain was found to be 15% increase in growth speed, the decrease in temperature sensitivity, and the improvement in cell survival rate.

Abstract:The effects on the growth characteristics of Heamatococcus lacustris of different ammonia concentrations（0.05~0.8 g/L） and different growth temperatures（22、25、28 ℃） and the effects of stress temperatures（30，35，38 ℃）on the astaxathin accumulation were studied in this paper. The main parameters OD，dry weight，chlorophyll a and astaxathin content were determined. The results show that Heamatococcus lacustris grows relatively better under lower concentration of ammonia nitrogen when using NH4Cl as nitrogen source. Its suitable growth conditions are as follows：temperature：22 ℃、NH4Cl：0.1 g/L，whereas the suitable stress temperature is 35 ℃.

Abstract:Amino acid nitrogen （AAN） is an important indicator representing the overall level of amino acids and low molecular weight peptides in the Chinese rice wine，which content can affect the quality and flavor of wine. The influence of post-fermentation process parameters to AAN generating were discussed in this work，which included the post-fermentation time and post-fermentation temperature by tracing the dynamic changes of protein content，AAN content and amounts of viable yeast cells， Tricine analysis by SDS-PAGE，and the physicochemical indexes and amino acid composition of fermentation broth. The results showed that the post-fermentation was the main period for AAN producing. The content of AAN in draft rice wine was increased 24.7%，31.7% and 37.6%，respectively when post-fermentation time prolonged from 14 days to 17 days， 20 days and 23 days. Yeast autolysis process offered a significant contribution to AAN at end of post-fermentation. Meanwhile it had positive effect on AAN formation when post-fermentation temperature raised at range of 13 ℃ to 17 ℃. The results implied that AAN was influenced by both activities of protein degradation enzymes and the degree of yeast autolysis when post-fermentation temperature was changed. Furthermore，the work indicated that amino acids are the main composition of nitrogen compounds，which nitrogen content accounted for more than 60% of AAN. This study is aimed to understand the effect of process conditions on the AAN generating，which will benefit the quality improvement of Chinese rice wine.

Abstract:In order to explore the specific monoclonal antibodies against Listeria monocytogenes for the immunological detection， irradiation life-style and thermal extinguishing method were used to produce Listeria nonocytogenes antibodies as the antigen，four hybridomas stably secreting McAbs to Listeria monocytogenes were developed. The results showed that fourhybridomas were successfully established，and the indirect ELISA titers of the ascites were 1：104～1：4×104. Their immune globulin subtypes were IgG1，Ig2a，Ig2b.Therefore，Listeria monocytogenes could be detected by a two-antibody-sandwich enzyme-linked immunosorbent assay （das-ELISA） formed by McAbs. The affinity constants （Kaffi） were from 1×107 L/mol to 1×1010 L/mol，which had no cross-reaction between the other compounds. The sensitivity of LM McAbs in the detection of antigen was as low as 1×103 CFU/mL. The high-titer，sensitive and specific anti-LM monoclonal antibodies were produced in this study，which makes it possible to develop immunoassay for detection of LM residues．

Abstract:One SOD high-producing strain was screened and identified from lactic acid bacteria. The SOD enzyme was purified. The microbial enrichment，Gram staining and Pyrogallol autoxidation determination method were used in the screening. Morphological characteristics，physiology and biochemistry experiments，16S-rDNA sequencing were used for the strain identification. The SOD enzyme was purified by ammonium sulfate precipitation，ultrafiltration，sephadex-G100，DEAE-Sepharose FF etc. A SOD high-producing strain （SD006） was obtained and identified as Lactobacillus plantrum. The activity of purified enzyme reached 37 959.2 U/g SD006 is a SOD high-producing probiotics strain of Lactobacillus plantrum. The SOD purified from Lactobacillus is valuable and can be developed and applied in the food industry.

Abstract:A sandwich immnuoassay based on monoclonal antibody for detection of Pseudomonas syringae pv. Maculicola was developed. Killed Pseudomonas syringae pv. Maculicola cell was used as immunogen to immunize BALB/c mice. The antisera was detected and the mice of highest antisera titer was selected for cell fusion. With hybirdoma technology，six cell lines secreting specific antibody to Pseudomonas syringae pv. Maculicola were obtained. Monoclonal antibodies （mAb） coded mAb1 and mAb6 were selected used as coating antibody and detection antibody based on pair-matching screening results. The sandiwich measure based on double antibodies were found to be high specific to targeted bacteria and no cross reactions to other bacteria including Burkholderia glumae，Xanthomonas oryzae pv. Oryzicola，Pantoea stewartii subsp.stewartii and Pseudomonas syringae pv.syringae. Under optimized conditions，the limit of detection was 1.5×105 cfu/mL and quantitative limit was 4.12×105 cfu/mL. This immunoassy was highly specific to Pseudomonas syringae pv. Maculicola and suitable for in-field detction，which provide a convient and fast tool in plant bacteria analysis.

Abstract:This study integrate enzymatic reaction with several membrane technologies to construct a method for comprehensive utilization of antioxidant peptide and other active substances from laver. The hydrolysate of laver was separated and purified through ultrafiltration（10 kDa cut-off molecular weight） and nanofiltration（500 Da cut-off molecular weight）. After optimizing the operation parameters of membrane separation，recovery rates of antioxidant peptide，polysaccharide，dietary fiber，and peptide reached 73.1%，55%，85%，and 18%，respectively. The peptides from laver showed the DPPH scavenging activity. The DPPH scavenging activity of 0.5 mg/mL nanofiltration peptide solution was up to 68.5%. Furthermore，the optimum pH value of laver antioxidant peptide was 8.5. Meanwhile，these peptides showed a good stability，it retained about 83% relative scavenging activity of DPPH radical after heated at 80 ℃ for 6 h.

Abstract:CGT⊿ECBMAmy gene from E. coli pET-20b（+）/CGT?驻E-CBMAmy was cloned and inserted into the expression plasmid pMA0911-SPs containing six different signal peptides（estA，bpr，vpr，yncM，yvgO and ywbN ），which were further transformed into B. subtilis WB600 for overexpression to obtain six recombinant strains. The results showed that estA was the optimal signal peptide among all the signal peptides under the same culture conditions，the yields of AA-2G was up to 0.81 g/L，obvious higher than that of the other five strains. Therefore，WBpMB/CGT?驻E-CBMAmy was chosen as the starting strain and the optimum conditions for extracellular expression of CGT?驻E-CBMAmy were investigated in shaking flask. Finally，the high expression of CGT?驻E-CBMAmy enzyme was achieved，and the yields of AA-2G was about 2.16 times higher than that before optimization and the conversion rate of VC reached 21.9%.

Abstract:The gene encoding collagenase from Pseudoalteromonas sp.NJ631 was cloned and expressed in E.coli cells. Subsequently，the bioactivity of recombinant protein，named PNJC，was assessed. The gene sequence encoding a putative collagenase，designed Pnjc，was obtained from the genome of Pseudoalteromonas sp. NJ631 using a genome mining approach. The Pnjc coding region was amplified and cloned into pET-28a Vector. The plasmids harboring the assembled full-length sequence encoding PNJC were transformed into the E.coli cells for the expression of the PNJC protein. The expression and purification of the recombinant protein were carried on according to manufacturer's instruction. Finally，the enzymatic activity of collagenase was assessed by using recombinant protein decomposing the type I collagen as substrate. The 1359-bp open reading frame with 45.62% GC was obtained from the genome of Pseudoalteromonas sp. NJ631，which was encoded of a protein of 452 amino acids and shared significant homology with the collagenase from the other genera. The PNJC was highly expressed by adding 0.2 mM IPTG. The assay of enzyme activity showed that the enzyme activity of PNJC was 160.34 U/mg，70.48% of the standard，confirming that the Pnjc gene encoded a functional collagenase. This is the first report of the cloning the expression of collagenase from marine bacteria Pseudoalteromonas sp. The PNJC encoded by Pnjc gene from NJ631 showed high bioactivity on decomposing the type I collagen，which are of potential value for the commercial exploitation.

Abstract:Based on results of the bioinformatics analysis，a mutant of recombinant human interleukin 29（rhIL-29mut33） was constructed by site-directed mutagenesis and megaprimer PCR. Then，it was successfully expressed in Pichiapastoris GS115. The rhIL-29mut33 showed strong anti-proliferation effects on liver cancer cell BEL7402，colon cancer cell HCT8，and gastric cancer cell SGC7901 at different concentrations，and all of them showed stronger than that of wild type rhIL-29. At a high concentration of the rhIL-29mut33，the inhibition ratios of the three tumor cells were （31.76±2.87）%，（22.47±0.26）%，and （34.41±0.40）%，respectively. The superior biological activities of rhIL-29mut33，especially the antitumor activity，make it have great potential applications in the drug industry.

Abstract:In order to identify the functional groups of peroxidase from the leaves of Ipmoea aquatica Forsk， peroxidas were isolated，purifiedandselectively modified by using BD，DTT，pCMB，Maleic anhydride，Chloramine-T，PMSF，NAI and BrAc separately and the change on activity were determined. The result showed that Arg，Lys，His and sulfhydryl involving in the composition of the enzyme active center should be considered as indispensable functional groups of peroxidase，while Met，Ser and Tyr residues should not be because they were not directly related to the activity of peroxidase from Ipomoea aquatica Forsk.

Abstract:Kombucha Fermentation conditions wereoptimized by pH value and sensory scores.，The fermentation substrates werebrown sugar，and the instant pu-erh tea：water was at 0.8 g∶1 000 mL.when brown sugar content was at 5.5 g and the pH value was at 3.7，the Kombucha beverage was orange，Pleasant aroma，moderate sweet and sour taste，also showed high membrane yield. The microbial results in optimization technique showed that Kombucha was composed of yeast，acetic acid bacteria and lactic acid bacteria. This study discovered new microorganisms which werebeneficial for Kombucha fermentation and human health.

Abstract:Ganoderma lucidum powder was extracted by ethanol solution of different concentration and the changes of the main functional components and antioxidant capacity of Ganoderma lucidum wine during extraction were studied. The results showed that the content of the total saponin， polysaccharides and protein were increased with time. The content of total saponin and total polysaccharides reached the peak at the 35 th day and then was declined slightly，and the content of total protein was declined after 30 days when it reached the peak. DPPH radical scavenging activity and ABTS radical scavenging activity reached the peak at the 35 th day. Positive correlation was found between antioxidant capacity and the content of total saponin，total polysaccharides and total protein. The antioxidant capacity and the content of functional components of Ganoderma lucidum wine varied with the concentration of ethanol solution used to extract Ganoderma lucidum for wine. The three main functional components could be extracted well with 70% ethanol solution，and the antioxidant capacity of the wine was the highest. So the best concentration of ethanol solution to extract Ganoderma lucidum for wine was 70%，and the best extraction time was about 35 days.

Abstract:The preliminary study on aroma compositions and fermented processes for Jerusalem artichoke wine was carried out using J. artichoke（Helianthus tuberosus L.） tubers as raw materials. Through comparing three kinds of yeasts on J. artichoke liquid fermentation experiments，the optimal yeast was selected. Then a single-factor and with two orthogonal-factor models were used to explore the optimal technology. Head Space Solid-Phase Micro-Extractions（HS-SPME） and Gas Chromatography-Mass Spectrometer（GC-MS） were used to determine the dominated aromas and flavor profiles of wine. The results were shown as follows：1） K.marxianus X2 was the best active inulinase.2） The optimal conditions were 30 ℃，5% inoculum dose，and 22 °Bx inulin when the yields of ethanol used as indexes，whereas the better conditions were 28 ℃，6% inoculum dose，and 21 °Bx inulin when the sensory quality of products as the scale index.3） Forty aromas and flavor components were characterized in J. artichoke wine，in which amyl acetates，ethyl hexanoates，and ethyl caprylates were dominated. A strain directly used for J. artichoke wine fermentation was elected to obtain the color，flavor and taste of J. artichoke wine.

Abstract:The contents of amylose，amylopectin and total starch were determined via Dual-wavelength spectrophotometry and Anthrone colorimetry in transgenic potato tubers，respectively. Furthermore，activities of GBSS，SSS were measured. Independent samples t test were performed by using statistical SPSS software. The results showed that the contents of total starch and amylopectin in transgenic potato tubers（K5） were significantly higher than that in control group，while activities of GBSS，SSS and the amylose content of K5 decreased significantly. This indicated that synthase gene of transgenic potato was affected by RNA interference in some extent.