引入CpG基序的汉滩病毒G2糖蛋白基因的克隆及表达

构建汉滩病毒G2糖蛋白的真核表达载体,并加入可增强小鼠免疫刺激作用的CpG基序,检测其可否在真核细胞中表达。参照Genebank中汉滩病毒M的全基因序列设计引物,引物两端引入可增强小鼠免疫刺激作用的CpG基序及双酶切位点,通过聚合酶链反应(PCR)获得含CpG基序的G2片段,并将其与T载体pMD18-T相连,测序后克隆至真核表达载体pcDNA3.1 上,将此真核表达载体以脂质体法转染至真核细胞Vero-E6中,利用间接免疫荧光法(IFA)检测发现转染后的Vero-E6中出现特异性的绿色荧光。结果表明本实验成功构建了汉滩病毒包膜糖蛋白G2的重组体。     

不同粒度羧甲基壳聚糖基复合抗菌剂抑菌性能的研究

为增加固体制剂的水溶性以及考查研磨后其抗菌性能的变化,制备了低银含量和低噻苯咪唑含量的羧甲基壳聚糖银噻苯咪唑（Ag—CMCTS-TBZ）复合抗菌剂,通过研磨和粒度分级将其分成C1（150—74μm）、C2（74—45m）和C3（-45μm）三种制剂,采用粒度分析、紫外、红外光谱及最小抑菌浓度（MIC）对其进行分析。研究表明,低含量银、噻苯咪唑的Ag—C／VICIS—TB均具有良好的水溶性,其溶解度都大于0．4％,其0．2％（w／v）的抗菌水溶液中的表观粒度在13—250nm之间;但在研磨分级之后,三种制剂中分子间的络合或者氢键键合状况发生了变化,而且导致其抗菌性能发生改变;其中C1对E．coli和S．aureus有较强的抑制,而其抑制A．niger的能力较弱;C3抗菌活性最低,对E．coli,S．aureus,C．albicans和A．niger的MIC值皆〉1000mg／L;Q的抗菌效果则介于其间。     

基于线粒体Cytb基因全序列探讨两爪鳖和山瑞鳖的系统进化关系

采用PCR方法对两爪鳖(Carettochelys insculpta)和山瑞鳖(Pdea steindachneri)mtDNA的细胞色素b基因(Cytb)进行了扩增和测序,并结合GenBank中已公布的其它15种龟鳖类和两种鳄类的同源序列,进行了序列变异比较和系统发生分析.以凯门鳄和扬子鳄为外群,采用邻接(neighbor ioining,NJ)法、最大简约(maximum likelihood,ML)法和最大似然(maximum parsimony,MP)法重建分子系统树.结果均支持将龟鳖目分为3支:侧颈龟科(Pelomedusidae)(非洲侧颈龟Pelomedusa subrufa),两爪鳖科(Carettochelyidae)(两爪鳖Carettochelys insculpta)和一支由其它15种现生龟鳘类组成的进化支.在这里两爪鳘科被认为可能代表一新的亚目;而山瑞鳖聚于鳖科(Trionychidae)中,与中华鳌(Pelodiscus sinensis)最先聚成一支,再依次与马来鳖(Dogania subplana)、非洲鳖(Trionyx triunguis)、印度箱鳖(Lissemys punctata)聚成一组.     

不同产地中华鳖的线粒体控制区序列分析及结构比较

采用PCR特异引物,扩增了两产地中华鳖（Pelodiscus sinensis）个体的mtDNA控制区（CR）及其邻近片段,测序获得了长度分别为1830bp和1630bp的序列。结合GenBank中已发表的韩国产中华鳖mtDNA的CR区序列,比较了3个产地中华鳖的CR区结构。分析显示：中华鳖不同产地mtDNA CR区DNA中的A＋T含量分别为60.5%、63.6%和64.8%,它们的5′、3′末端以及CSB1-CSB2之间均存在丰富的可变数目串联重复序列（variable numbers oftandem repeats,VNTR）。基于mtDNA CR区序列和结构分析,显示中华鳖不同产地的野生个体中存在丰富的遗传多样性。     

芦蒿不同浸提物化感活性的比较研究

采用水、甲醇、乙醇、丙酮和乙酸乙酯等不同溶剂对整株芦蒿进行浸提,并用生菜和紫花苜蓿作为受试植物,对不同浸提物进行了化感活性的比较研究。5种不同溶剂的芦蒿浸提物对种子的萌发率和幼苗生长均表现出化感潜势,与对照组相比,0.005 g/mL的芦蒿甲醇提取物对生菜和紫花苜蓿种子的萌发具有轻微的促进作用,当浓度达到0.04 g/mL以上时,对生菜和紫花苜蓿种子的萌发和幼苗生长均具有不同程度的抑制作用（p〈0.05）。不同浓度的提取液对植物的化感作用强度不同,对种子萌发率、根长、芽长均有不同程度的影响。根据比较,甲醇溶剂提取物的化感活性最强,其是深入研究芦蒿化感物质有效的提取剂。     

人胎盘滋养层细胞原代的体外培养与改进

目的:建立与改进纯度较高的适于实验研究的人绒毛膜滋养层细胞。方法:采用胰蛋白酶消化法消化人正常妊娠6～8周胎盘组织,以35%、45%2个Percoll密度梯度进行分离纯化,并用免疫组化及透射电镜等技术对其生物学特性、细胞内部结构进行观察。结果:胎盘组织中滋养层细胞角蛋白染色阳性,血管内皮细胞及基质成分波形蛋白染色阳性,经该法分离纯化的细胞角蛋白染色阳性者(滋养层细胞)占90%以上,透射电镜观察示所获细胞有典型滋养层细胞结构。结论:该法简便易行,可获得合乎实验要求的人滋养层细胞,可供后续实验研究。     

Molecular analysis of the bacterial community in a continental high-temperature and water-flooded petroleum reservoir

Water from a continental high-temperature, long-term water-flooded petroleum reservoir in Huabei Oilfield in China was analysed for its bacterial community and diversity. The bacteria were characterized by their 16S rRNA genes. A 16S rRNA gene clone library was constructed from the community DNA, and using restriction fragment length polymorphism analysis, 337 randomly selected clones were clustered with 74 operational taxonomic units. Sequencing and phylogenetic analyses showed that the screened clones were affiliated with Gammaproteobacteria (85.7%), Thermotogales (6.8%), Epsilonproteobacteria (2.4%), low-G+C Gram-positive (2.1%), high-G+C Gram-positive, Betaproteobacteria and Nitrospira (each <1.0%). Thermopilic bacteria were found in the high-temperature water from the flooded petroleum reservoir, as well as mesophilic bacteria such as Pseudomonas-like clones. The mesophilic bacteria were probably introduced into the reservoir as it was being exploited. This work provides significant information on the structure of bacterial communities in high-temperature, long-term water-flooded petroleum reservoirs.     

Analysis of alkane-dependent methanogenic community derived from production water of a high-temperature petroleum reservoir

Microbial assemblage in an n-alkanes-dependent thermophilic methanogenic enrichment cultures derived from production waters of a high-temperature petroleum reservoir was investigated in this study. Substantially higher amounts of methane were generated from the enrichment cultures incubated at 55 °C for 528 days with a mixture of long-chain n-alkanes (C₁₅–C₂₀). Stoichiometric estimation showed that alkanes-dependent methanogenesis accounted for about 19.8% of the total amount of methane expected. Hydrogen was occasionally detected together with methane in the gas phase of the cultures. Chemical analysis of the liquid cultures resulted only in low concentrations of acetate and formate. Phylogenetic analysis of the enrichment revealed the presence of several bacterial taxa related to Firmicutes, Thermodesulfobiaceae, Thermotogaceae, Nitrospiraceae, Dictyoglomaceae, Candidate division OP8 and others without close cultured representatives, and Archaea predominantly related to uncultured members in the order Archaeoglobales and CO₂-reducing methanogens. Screening of genomic DNA retrieved from the alkanes-amended enrichment cultures also suggested the presence of new alkylsuccinate synthase alpha-subunit (assA) homologues. These findings suggest the presence of poorly characterized (putative) anaerobic n-alkanes degraders in the thermophilic methanogenic enrichment cultures. Our results indicate that methanogenesis of alkanes under thermophilic condition is likely to proceed via syntrophic acetate and/or formate oxidation linked with hydrogenotrophic methanogenesis.     

刀鲚幼鱼消化系统的组织形态学结构

采用光镜和扫描电镜观察长江刀鲚(Coilia nasus)幼鱼消化系统组织形态学结构。结果显示,刀鲚体长,口裂大,含有犬齿状的颌齿和尖锥状的腭齿,具有5对鳃弓,鳃耙长度明显大于鳃丝且表面附着不规则绒毛状细齿;胃呈"Y"型,胃与肠连接处具有16～21个指状环形幽门盲囊;肠为直肠,较短,比肠长为0.241±0.080;肝分为两叶,胰为独立的器官。刀鲚口咽腔为复层鳞状上皮,含有腺体、大量椭圆形黏液细胞、少量杯状细胞及味蕾;胃黏膜都为典型的单层柱状上皮,含有较多由上皮凹陷形成的胃小凹和胃腺;幽门盲囊具有20～25个丰富的褶皱,占满大部分幽门盲囊腔,黏膜层具有微绒毛;中肠黏膜上皮最发达,形成的褶皱细长且连接成网状,单层柱状上皮与复层扁平上皮交替分布。观察结果表明,刀鲚消化系统具有典型肉食性鱼类特征。