A yeast strain MJ2 that was found to produce a higher amount of γ-aminobutyric acid (GABA) was isolated from the surface of kiwi. Phylogenetic analysis based on the ITS sequence and morphological, biochemical studies indicated that it may belong to Saccharomyces cerevisiae . Under optimum conditions in Czapek’s broth medium with 0.5 % monosodium glutamate, it produced GABA at a concentration of 5.823 g/L after 48 h. A full-length glutamate decarboxylase gene ( Scgad ) was cloned by PCR amplification. The open reading frame (ORF) of the Scgad gene was composed of 1,755 nucleotides and encoded a protein (585 amino acids) with a predicted molecular weight of 65.897 kDa. The deduced amino acids sequence of Scgad shows 100 %, 65 % and 62 % similarity with S. cerevisiae , Candida glabrata and Kluyveromyces lactis GAD in the polypeptide level, respectively. The Scgad gene was expressed in Escherichia coli BL21 (DE3) cells, and the expression was confirmed by Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE) analysis. The results suggested that the S. cerevisiae GAD ( Sc GAD) was successfully encoded in E. coli BL21 (DE3) cells. Furthermore, the enzyme activity of Sc GAD encoded in E. coli BL21 (DE3) had been significantly enhanced using artificial neural network linked with genetic algorithm (ANN-GA) method.

538 transformants from T-DNA mutant library of Monascus purpureus mediated by Agrobacterrium tumefaciens were used as materials,9 respresentative transformants with significant changes were selected through the colony and mirco-morphological observation,and verified by PCR amplification.The results showed that there were T-DNA insertions in transformants genome.Through stability examination,all 9 respresentative transformants could be steadily inherited.Based on this,two main metabolites of 9 respresentative transformants,which were pigments and γ-aminobutyric acid(GABA),were analyzed respectively by using the methods of ethanol extraction and paper chromatography.The results showed that the capacity of producing metabolites by transformants changed in different degrees compared with the original strain.It provided the foundation for the regulation mechanism of Monascus metabolite research.

The results of bacteriostasis experiments demonstrated that strain ACT-2 suppressed a wide range of bacterial pathogens in vitro,while no obvious antimicrobial activities towards fungal pathogens.Antibacterial stability of Streptomyces termitum strain ACT-2 fermentation broth was studied while Xanthomonas oryzae pv.oryzae physiological strain P6 was chosen as indicator bacteria.The fermentation broth could show obvious bacteriostatic activity when the tempreture was below 80 ℃ or exposured in the 4 000 lx illuminant;the diameter of inhibition zone decreased when the tempreture came up to 90 ℃.The fermentation broth was sensitive to acidity and alkalinity,the activity decreased after treatment.

Two main phytotoxic and antifungal phthalic acid butyl isobutyl ester (1) and radicinin (2) were isolated from the culture of Curvularia sp. FH01, a fungus residing in the Atractomorpha sinensis gut. The structures of isolated metabolites were established on the basis of spectral analysis. Metabolites 1 and 2 exhibited significant phytotoxic activity against the radical growth of Echinochloa crusgalli with their IC(50) values of 61.9 and 5.9 μg/mL, respectively, which were comparable to that 2,4-dichlorophenoxyacetic acid (2.0 μg/mL) used as a positive control. The antifungal test results showed that compound 2 possessed strong antifungal activity against Magnaporthe grisea (IC(50)=16.3 μg/mL) and Valsa mali (IC(50)=18.2 μg/mL). The findings of the present study suggest that bioactive properties of the fungus FH01 can be attributed to its major components, phthalic acid butyl isobutyl ester and radicinin, and both agents have a potential to be used as herbicide and fungicide.

从浙江省金华市郊区水稻试验田土壤筛选获得1株抗白叶枯病菌的放线菌ACT-2菌株.经形态学、16S rDNA序列分析和进化树建树分析,鉴定为白蚁链霉菌(Streptomyces termitum).生理生化实验显示,ACT-2菌株具有较强的环境适应能力.抗菌谱实验表明,ACT-2菌株对细菌类病原体具有较强的拮抗能力,对真菌类病原体效果不明显.ACT-2菌株的菌体和发酵液均对白叶枯病菌表现出明显的拮抗能力,抑菌圈直径分别达到38.00 mm和21.73mm.经发酵条件优化,ACT-2菌株在培养温度为28℃、初始pH7.5、摇床转速为180r/min、装液量为50 mL/250 mL,发酵液抑菌效果达到最大,抑菌直径达到26.20 mm.

The separation process of γ-Aminobutyric acid(GABA) from monascus fermentation broth was researched.Three types of decolorants were selected,and the D101 exhibited high decoloring ratio and high GABA yield.The static and dynamic adsorption-desorption performances were investigated to obtain optimized conditions of GABA separation by D001 ion-exchange resins.The result showed that the optimum parameters were as follows:original pH of decolor fermentation broth,sample flow-rate of 1 BV/h,elution method of two steps by using water and 2 mol/L aqueous ammonia.The transparence needle crystal of GABA was obtained after the effluent concentrated.The total recovery rate was 45.4%.

从浙江省金华市花生地土样中筛选、分离纯化得到1株产聚-β-羟基丁酸编号为PX-95的芽胞杆菌,PHB产量为135.81 mg/L。根据形态特征、生理生化特征初步鉴定为芽胞杆菌属蜡样芽胞杆菌群,16S rDNA序列分析显示PX-95菌株与苏云金芽胞杆菌以及蜡样芽胞杆菌均具有高度同源性,最后采用扩增gyrB（DNA促旋酶B亚单位）基因的方法将其鉴定为苏云金芽胞杆菌（Bacillus thuringiensis）。

红曲霉(Monacus purpureus )是我国重要的传统食用和药用微生物,能分泌红曲色素、莫纳卡琳(Monacolin K)、γ-氨基丁酸、麦角固醇等物质,被广连用于生产食品发酵剂、食品着色剂、保健食品和药品等,在我国有上千年的应用历史.本实验以根癌农杆菌(Agrobacterrium tumefaciens)介导的红曲霉突变体库中754株突变子菌株为材料,通过乙醇提取法筛选出了9株产红曲色素发生显著变化的色素突变子,其中S50在505 nm波长处的色价降为原始菌株S的0.12倍,几乎变为白化株.300 ～ 600 nm波长下UV-VIS扫描图谱特征显示,色素突变子不仅色价发生了变化,而且吸收波长、吸收峰值也发生了不同程度的变化.最后,对色素突变子菌落形态、显微结构、潮霉素抗性、遗传稳定性以及产桔霉素能力等进行分析,为研究红曲霉产色素相关基因奠定了基础.

采用响应面分析法对草酸青霉（Penicillium oxalicum）L5菌株液体发酵产果胶酶条件进行了优化。结果表明:桔皮粉、米糠及硫酸铵的添加量分别为4.85%、5.89%、0.97%,摇瓶初始pH为6.0~8.0,接种量为9%时,优化后的果胶酶活达54 391.70 U,是初始酶活18 148.00 U的3倍。另外,对其果胶酶性质进行了初步探讨,结果表明:该酶较适反应温度和pH分别为50℃和5.0;30~50℃时有较好的热稳定性,pH值为5.0时稳定性最佳。

The extraction conditions for producing exopolysaccharide by Monascus purpureus MP-66 were optimized in order to enhance the extraction yield.Single-factor experiments were carried to investigate the influences of the alcohol volume fraction,the sedimentation time of alcohol,and the pH value of fermentation broth on extraction yield of Monascus exopolysaccharide.On the basis of single-factor experiments,a quadratic polynomial regression equation forecasting model was set up by designing a Box-Behnken central composite experiment involving of three factors and three levels and the method of response surface analysis,thus the optimized extraction conditions for Monascus exopolysaccharide were obtained.According to the results,alcohol sedimentation time was 16.5 h,the alcohol column fraction was 80%,and the pH value of fermentation broth was 8.05.Based on these conditions,the yield of Monascus exopolysaccharide could reach 7.40 g/L.

A red pigment producing bacterial strain was screened from a freshwater fish body through isolating color bacteria in nature.This strain was identified as Serratia marcescens based on morphological,physiological characters and 16S rDNA gene sequence.This pigment was demonstrated belong to prodigiosin on the basis of UV spectrum and TLC.The yield of crude prodigiosin reached at 475 mg/L.

A high-yield pectinase strain L5 was screened from sludge.It was identified as Penicillium oxalicum by morphological observation,and internal transcribed spacer(ITS) sequence analysis.This strain was cultured at the condition of 28 ℃,pH 6.0,160 r/min for 70 h,its pectinase activity reached to 39 940 U·mL-1·min-1.

通过优化各种转化因素,建立了根癌农杆菌(Agrobacterium tumefaciens)介导红曲霉(Monascus)的高效转化体系:红曲霉在PDA培养基培养21 d后收集孢子,制备红曲霉孢子悬浮液,浓度为106 个/mL,根癌农杆菌浓度为OD600值0.5,诱导剂AS浓度为100 μmol/L,农杆菌与红曲霉在25 ℃共培养3 d.采用此转化体系构建了含有530多个转化子的红曲霉T-DNA插入突变体库.随机选取50株转化子菌株进行分子验证和稳定性检测,证明T-DNA成功插入红曲霉基因组DNA中,并能稳定遗传.最后,通过形态观察筛选出8株变异较大的菌株,为以后的红曲霉基因功能研究奠定了一定的基础.

从8个正向引物和11个反向引物随机组合成的88个引物对中筛选出14个引物组合,对19株酵母菌进行PCR扩增。采用正交试验设计方法,对SRAP的影响因素进行研究,从Mg2+、dNTPs、DNA浓度、引物、TaqDNA聚合酶设计5因素4水平进行优化试验,以确定PCR电泳分析的最佳条件。14个引物组合共扩增出279条带,其中92.5%呈多态性,相似系数范围为0.60~1.00。

In the present paper, Fourier transform infrared spectrometer was applied to gain the infrared spectra of Hypnum fertile Sendtn. samples with different concentration of Cd2+ for 7 days to study the changes in chemical components, and some physiological characteristics such as malondialdehyde (MDA), solubility sugar (SS) and solubility protein (Pr) were also studied. Malondialdehyde and solubility sugar were measured by anthrone colorimetry and solubility protein was measured by brilliant blue and commassie dye. The results showed that with the enhancement of Cd2+ concentration, the contents of solubility sugar and solubility protein were enhanced at first and then decreased. However, it was just the opposite for the content of malondialdehyde. Based on the indices of wave number-absorbance, the infrared spectra of samples with different concentration of Cd2+ were compared in terms of changes in the physiological characteristics. The two bands at 2 920 and 2 855 cm(-1) in the CH stretching region (3 000-2 800 cm(-1)) of the spectra denote the asymmetric and symmetric lipid CH2 stretching vibrations, whose band height's variety was related to the change in malondialdehyde content in plant. The distinctive amide-I band at 1 635 cm(-1) and amide-II band at 1 545 cm(-1) arise from endogenous proteins. In the region below 1 500 cm(-1), a variety of characteristic IR group frequencies can be observed that are difficult to assign but the shape of the spectra in this region is characteristic and can be used as a "finger print" of the tissue. The band at 1 036 cm(-1)-C-O, -C-C or CH stretching region of the spectra denotes the carbohydrate related to the content of solubility sugar. The results showed that there was good correspondence between the changes in the physiological characteristics and the changes of samples based on the indices of wave number-absorbance of FTIR spectra, and the FTIR was more sensitive and convenient. Therefore, Fourier transform infrared spectrometry can be accepted as a fast and effective method to study bryophytes for monitoring the pollution degree of heavy metal in environment.

Experiment was carried out on influence of copper stress on physiological and biochemical characteristics of Hypnum fertile and Anoectangium aestivum by indoor simulated culture. The results indicated that under the stress of copper, protective enzymes and elements closely related to the adversity of H. fertile and An. aestivum changed with evident relation of dosage/efficiency and duration/efficiency. Chlorophyll content and the activity of peroxidase (POD) increased first then decreased with the increase of concentration and duration of stress, while proline(Pro) content and malondialdehyde (MDA) content increased continually. Changes of physiological indexes of H. fertile were more evident under stress of copper compared with that of An. Aestivum, meaning that H. fertile was more sensitive to copper stress, and could be indicator for copper pollution.