一株海洋红法夫酵母Phaffia rhodozyma RP-306产虾青素的发酵条件优化

为提高海洋红法夫酵母Phaffia rhodozyma RP-306的虾青素产量,本研究利用单因素和响应面实验对其发酵条件进行了优化。首先用单因素实验确定了蔗糖、酵母粉、硫酸铵、pH、温度和装液量6个因素的初始值,再利用Plackett-Burman实验筛选出了影响较大的3个因素:蔗糖、pH、温度,并采用最陡爬坡实验确定了因素的合理范围,最后采用响应面法中的中心组合设计进行优化,其优化发酵条件为:蔗糖29.93 g/L,酵母粉2.0 g/L,硫酸铵5.0 g/L,pH 5.27,温度19.9℃。在此条件下虾青素产量从13.46 mg/L提高到17.04 mg/L,比优化前提高了26.61%。以优化后发酵条件进行分批发酵,虾青素产量达到28.86 mg/L,为进一步发酵放大研究奠定了良好基础。     

Bacterial community in sediment from the Western Pacific "Warm Pool" and its relationship to environment

Total DNAs were extracted from different sections of deep sea sediment core sample collected from the Western Pacific "Warm Pool". The bacterial 16S ribosomal DNA (rDNA) clone libraries were constructed and analyzed by PCR-restriction fragment length polymorphism (RFLP) and DNA sequencing. The bacterial communities in these samples and their relationship to environment were analyzed consequently. The results indicated that among eight main bacterial groups found in these sediments, members of the γ-Proteobacteria were most abundant in each section of sediment core sample and the genus Colwellia belonging to γ-Proteobacteria was dominant in this area. Members of the α-Proteobacteria were found commonly existing in these samples, while members belonging to β-Proteobacteria were seldom detected. The diversity of bacterial communities from different sections of sediment core sample was δ- and ε-Proteo-bacteria and the bacterial group including genera Cytopahga, Flexibacteria and Bacteroides (CFB group)     

深海羽毛降解菌Bacillus sp. JM7极端碱耐受角蛋白酶的酶活性质研究

Bacillus sp. JM7, a strain isolated from the deep-sea of the South China Sea, was found to efficiently degrade 79.4% native chicken feather within 30 h. Scanning electron microscopy analysis showed that JM7 strain could gradually degrade feather by modifying the microstructure of feather keratin. A total of 25 protease genes were predicted from the draft genome of JM7 strain, among which a predicted subtilisin-like serine protease(designated as Ker02562) was further characterized for its keratinolytic activity. The recombinant Ker02562 functioned at a wide range of temperatures from 30℃ to 60℃, with an optimum at 40–50℃. Ker02562 was highly active at various pHs ranging from 5.0 to 13.0, with a maximum activity observed at pH 7.0–9.0. Remarkably, recombinant Ker02562 was stable in extreme alkaline environments(pH 10–13), which was much better than most other reported keratinases. Collectively, these favorable properties could make Bacillus sp. JM7 and Ker02562 attractive to be applied in the detergent formulation and feather bioconversion.     

深海噬菌体BVE2内溶素的重组表达及其活性研究

本研究从一株来源于西南印度洋沉积物的噬菌体BVE2基因组中得到了一条全长702 bp,编码噬菌体内溶素的基因Lysin132,编码的蛋白共含有234个氨基酸残基,预计分子量为25.74 kDa;氨基酸序列同源比对结果表明BVE2 Lysin132具有多个酰胺酶活性位点。进化树分析结果表明BVE2 Lysin132是一种N-乙酰胞壁酰-L-丙氨酸酰胺酶。将Lysin132克隆到pEASY-Blunt E2 Expression Vector中,构建了E.coli-pEASY-Lysin132表达菌株。成功用IPTG诱导表达,得到了大量带有6×His标签的重组BVE2 Lysin132。经镍柱纯化后进行SDS-PAGE电泳,得到单一目的蛋白条带。酶学性质分析结果表明,重组BVE2 Lysin132的最适反应温度为30℃,在中高温下热稳定性良好,在10～50℃下孵育90 min,仍能保持70%以上的酶活力;最适pH为7.0,在pH 6.0～8.0时均能保持80%以上的酶活力,表明该酶有较宽的pH作用范围;Na⁺、Mg²⁺可以使酶活力显著提高20%以...     

一种诱导模式和甘油补料优化策略提高重组大肠杆菌BL21（DE3）β-琼胶酶的产量

琼胶酶是作用于琼胶的水解酶,在食品、化妆品和制药工业中有广泛的应用。本文建立了一种基于诱导模式和甘油补料优化的高细胞密度和高产β-琼胶酶策略,同时可以较好的控制乙酸盐产量。首先,在诱导前期采用不同的比生长速率（μ）的甘油指数补料策略。结果表明,低的比生长速率（μ=0.2）是细胞生长和β-琼胶酶产生的最佳条件。其次,研究了诱导阶段诱导温度和诱导物浓度对细胞生长和β-琼胶酶产生的影响。当异丙基-β-d-硫代半乳糖苷（IPTG）诱导时,在20℃下0.8mmol/L的IPTG诱导策略对β-琼胶酶的产生效果最佳。采用1.0g/（L·h）的乳糖连续补料策略诱导培养,β-琼胶酶活性达到112.5U/mL,是目前报道的产量最高的β-琼胶酶。此外,β-琼胶酶能直接酶解龙须菜粉,产生新琼寡糖,水解产物为新琼胶四糖（NA4）和新琼胶六糖（NA6）。本文的研究为β-琼胶酶的工业化生产和应用提供了较好的理论依据。     

数值流形方法中覆盖位移函数的改进

通过对数值流形方法刚度矩阵形成的研究,提出一种改进的覆盖位移函数,以改善刚度矩阵计算。分析表明,改进的覆盖位移函数可使刚度矩阵的局部大数数量级明显的降低,提高计算效率,给矩阵求解提供适当的意见,并给出算例验证方法的正确性。     

我国近海中度嗜盐菌的分离筛选及其产酶多样性分析

为了解我国近海中度嗜盐菌系统发育多样性及产酶特性,从天津、山东、江苏、福建、海南等地近海采集样品为研究对象,分离筛选得到108株中度嗜盐菌,其中有26株至少产一种酶。通过对其形态特征、生理生化、16SrRNA基因序列及系统进化树进行分析,将26株菌鉴定为细菌域的Halomonas、Idiomarina、Virgibacillus、Pontibacillus、Oceanobacillus、Halobacillus和Marinilactibacillus属的菌株。它们与相应属的模式菌株16SrRNA基因序列相似性在97%~100%不等,其中有些菌株可能代表不同的分类单元。底物特异性试验表明,分离的26株中度嗜盐菌13株产蛋白酶,19株产淀粉酶,13株产酯酶,4株产纤维素酶。其中6株产3种酶,11株产2种酶。研究结果表明中度嗜盐菌具有系统发育和产酶多样性,同时蕴藏着较多新的微生物类群,为嗜盐微生物资源应用开发提供了参考。