共表达VP3与hIL-18基因真核重组质粒的构建及对人结肠癌细胞的影响

先将鸡贫血病毒VP3基因插入真核表达载体pVAXl的多克隆位点，再将pVAX1载体上的IRES启动子及新霉素基因一同切下插入pVAXl的VP3基因下游，然后切下新霉素基因，将pIRES1基因插入其中，构建成pVVP3IL-18，将pVVP3IL-18转染Heda细胞。间接免疫荧光表明，在细胞膜和细胞浆观察到了特异的黄绿色荧光，证明表达了hIL-18。pVVP3IL-18转染人结肠癌细胞HCY-116后，经AO／EB染色及电镜观察，可见大量细胞凋亡。说明真核重组质粒pVVP3IL-18可在HeLa细胞中表达，并可在体外诱导人结肠癌细胞HCT-116凋亡。     

不同浓度Mg-6Zn合金浸提液对肠上皮细胞凋亡及其相关基因Caspase-3表达的影响

研究不同浓度Mg-6Zn合金浸提液对大鼠肠上皮细胞IEC-6的凋亡及凋亡相关基因Caspase-3表达的影响。将Mg-6Zn合金制成不同浓度(20%和40%)的浸提液体外培养大鼠肠上皮细胞IEC-6,并以含10%胎牛血清的高糖DMEM(Dulbecco′s modified Eagle′s medium)培养基作为阴性对照。采用噻唑蓝(MTT)法通过计算细胞相对增殖率检测不同浓度合金浸提液对IEC-6细胞的毒性作用;采用流式细胞仪检测不同浓度合金浸提液对IEC-6细胞凋亡的影响;采用逆转录聚合酶链反应(RT-PCR)法检测不同浓度合金浸提液对IEC-6细胞凋亡相关基因Caspase-3mRNA表达的影响。实验结果表明,20%和40%两种浓度的镁合金浸提液在体外培养IEC-6细胞第七天时细胞毒性分级分别为0级和1级,属于无毒性,符合医用材料细胞毒性分级的合格标准。40%和20%浓度镁合金浸提液中,IEC-6细胞的凋亡比例及Caspase-3活化度相对于对照组都增高,差异有统计学意义(P0.05),40%浓度组中,IEC-6细胞的凋亡比例及Caspase-3活化度相对于20%浓度组增高,差异有统计学意义(P0.05),说明Mg-6Zn合金浸提液在一定范围内处于较高浓度时易诱导IEC-6细胞的凋亡及Caspase-3的表达,且随浓度升高影响越大。     

壳寡糖通过降低线粒体膜电位促进A549细胞凋亡

壳寡糖(chitooligosaccharides,COS)分子量小,可溶性好,具有抗肿瘤等多种生物活性.以人肺腺癌A549细胞系为研究对象,探究COS对A549细胞线粒体膜电位、细胞生长与死亡的影响.MTT结果表明COS浓度越高,刺激时间越长,其对A549细胞生长的抑制作用越明显;流式细胞仪检测结果显示,1mg/mL的COS明显地降低了细胞线粒体膜电位水平;活死细胞荧光染色实验表明,COS组死亡细胞比率提高,与对照组相比具有显著性差异.结果提示,COS可能通过降低线粒体的膜电位而促进A549细胞凋亡.     

重组质粒pcFlaA在青石斑鱼肌肉中的表达研究

在哈维氏弧菌TS-628菌株鞭毛丝蛋白FlaA基因末端加上一段编码Flag短肽的核苷酸序列作为检测标记后,将该基因克隆到真核表达载体pcDNA3.1( ),酶切、PCR扩增及重组质粒测序证实基因片段插入正确,将该重组质粒命名为pcFlaA.将pcFlaA以肌肉注射方式免疫青石斑鱼.免疫后第7天开始检测鞭毛丝蛋白在石斑鱼肌肉中的表达状况,之后每隔1周检测1次,共检测4次.首先采用PCR技术在DNA水平检测重组质粒转染石斑鱼肌肉细胞的情况,再以RT-PCR法在mRNA水平上检测转染质粒在鱼肌肉中的转录,最后以免疫组化染色技术在蛋白质水平上检测目的蛋白的表达.结果在DNA及mRNA水平上均可检测到目的条带,在蛋白质水平上可检测到明显阳性位点,由此证实pcFlaA可以转染石斑鱼肌肉细胞并可在其中进行表达,而且质粒在鱼体内持续表达的时间至少1个月.     

羟基磷灰石纳米颗粒诱导巨噬细胞凋亡及其与HSP70相互关系的研究

研究羟基磷灰石纳米颗粒(HAp nanoparticles)引起巨噬细胞凋亡,及其与热休克蛋白Hsp70的相关性.THP-1单核细胞株经PMA诱导成为巨噬细胞,分别加入0、20、100μg/ml HAp纳米颗粒,培养24h后经PI和AnexinV染色,在荧光显微镜下观察,用流式细胞仪检测细胞的凋亡情况,应用western blot 方法检测Hsp70蛋白分子表达的改变.显微镜观察显示THP-1单核细胞株能被诱导成巨噬细胞.荧光显微镜显示20、100μg/ml HAp纳米颗粒组凋亡的阳性信号明显强于0μg/ml对照组;流式细胞仪检测结果也证实HAp纳米颗粒组比对照组凋亡率明显增高(P1＜0.0001,P2＜0.0001),且凋亡的程度呈浓度依赖性(P3=0.025).Western Blot 检测显示20、100μg/ml HAp纳米颗粒组较对照组Hsp70蛋白表达增加,并且随颗粒浓度增高而表达增加.     

扇贝多肽(PCF)抑制紫外线诱导的HaCaT细胞凋亡

建立了紫外线A和B辐射诱导的永生化角质形成细胞(HaCaT)凋亡的模型,采用四甲基偶氮唑盐法、琼脂糖凝胶电泳、流氏细胞仪、酶化学方法和蛋白印迹等技术,研究了扇贝多肽(PCF)抑制紫外线诱导HaCaT细胞凋亡的分子机制。实验发现,PCF可明显抑制紫外线A和B诱导的HaCaT细胞凋亡。1.42~5.68mM剂量范围内PCF剂量依赖性抑制紫外线诱导的HaCaT细胞内活性氧的生成,并能提高细胞内抗氧化酶活性,增强细胞总抗氧化能力,减少脂质过氧化产物的产生;同时,PCF也抑制细胞内JNK蛋白的磷酸化及Caspases-3蛋白的活化。通过加入ROS、JNK和Caspase-3抑制剂证实,PCF作为抗氧化剂减少了细胞内ROS的生成并提高了细胞内抗氧化酶活性,进而阻断了ROS→JNK→Cas-pase-3信号级联反应,抑制了紫外线辐射诱导的HaCaT细胞凋亡。     

中国明对虾线粒体MnSOD在大肠杆菌中的重组表达、产物纯化及活性测定

采用PCR方法扩增编码中国明对虾线粒体锰超氧化物歧化酶(MnSOD)成熟肽的基因,并克隆到大肠杆菌表达载体pCRR T7/NT TOPOR TA中进行体外重组表达.重组质粒转化大肠杆菌BL21 (DE3) pLysS后,经IPTG诱导表达产生包涵体形式的目的蛋白.对重组蛋白进行 LC-ESI-MS 分析的结果表明,融合蛋白的三个肽段与中国明对虾线粒体MnSOD 相应肽段完全一致.将重组蛋白通过金属螯合柱进行纯化,进而透析、复性,最后获得了活性高达2373U/mg的重组蛋白.中国明对虾线粒体MnSOD的成功表达,为深入研究其在中国明对虾免疫反应和抗氧化胁迫中的作用奠定了基础.     

As2O3纳米粒的制备和体外治疗肺癌及其抗转移机制实验

采用改良的溶胶-凝胶法制备系列的As2O3纳米粒,用透射电镜、扫描电镜、能谱仪、图像分析系统等进行表征及特性检测．应用MTT法研究As2O3纳米粒体外对细胞的增殖抑制作用;用流式细胞仪测定As2O3纳米粒及亚砷酸诱导细胞的凋亡率;用免疫组织化学半定量法检测As2O3纳米粒及亚砷酸处理细胞后Bcl-2、Bax、CD44v6和P53基因的表达改变．研究结果表明,制备的As2O3纳米粒在电镜下呈圆形或椭圆形,分散性较好,平均直径约为80nm、110nm、130nm、150nm和450nm;体外细胞实验证实As2O3纳米粒抗肺癌A-549细胞的效应强于亚砷酸溶液;免疫组织化学半定量法显示As2O3纳米粒有较强的诱导肺癌细胞凋亡的作用,可能与其改变Bcl-2和Bax基因表达（Bcl-2／Bax比值降低）及促进P53基因的表达、抑制CD44v6基因表达有关．     

几种齿科合金生物相容性的比较

用银钯合金、镍铬合金、钴铬合金、2号合金的浸提液体外培养小鼠成纤维细胞(L929),以含10%胎牛血清的RPMI1640培养基作为阴性对照组。用5种浸提液分别培养L929细胞4 h、7 h、24 h、48 h后,用MTT法检测细胞增值率并计算caspase-3活化度。在荧光显微镜下观察48 h后各组细胞的状态。使用凋亡相关因子caspase-3的表达强度辅以时间梯度分析其与齿科合金生物相容性的相关性。结果表明:加样培养48 h后各实验组均有大量凋亡细胞,但是有明显的不同,在镍铬合金组内可见橙色坏死细胞;各组细胞的毒性为0级,其细胞增值率的排序为:钴铬合金组>2号合金组>银钯合金组>镍铬合金组;各实验组的Caspase-3活性有显著的差异(P<0.01),Caspase-3活化度由高到低的排序为:钴铬合金组>2号合金组银钯合金组>镍铬合金组。各时间点Caspase-3活化度也有显著的差异(P<0.01),其由大到小的排序为:4 h、48 h、7 h、24 h;这几种齿科合金生物相容性由高至低的排序为:钴铬合金>2号合金>银钯合金>镍铬合金。镍离子、银离子是影响其生物相容性的主要因素。在时间梯度的辅助下评价4种齿科合金生物相容性,Caspase-3与MTT的结果一致.     

牙鲆淋巴囊肿病毒主要衣壳蛋白基因片段真核表达载体的构建、表达与免疫效果评价

用淋巴囊肿病毒LCDV-cn感染牙鲆鳃细胞系FG-9307,提取细胞总RNA,用RTPCR法获得了LCDV-cn主要衣壳蛋白(MCP)0.6kb基因片段.将该LCDV-cn-MCP0.6kb片断克隆入真核表达载体pEGFP-N2,得到重组质粒pEGFP-N2-LCDV-cn-MCP 0.6.采用脂质体法将重组质粒转染入牙鲆鳃细胞系FEC,并进行瞬时表达.通过荧光显微镜观察和特异性RT-PCR检测,证实重组质粒pEGFP-N2-LCDV-cn-MCP 0.6kb已成功转染到FEC细胞,并得到了初步表达.将重组质粒肌注入牙鲆体内,检测牙鲆外周血、肠、脾脏、前肾和淋巴细胞的增殖反应、呼吸爆发活性及抗体产生水平.结果表明,构建的核酸疫苗pEGFP-N2-LCDV-cn-MCP 0.6可诱导牙鲆特异性体液免疫和细胞免疫,具有明显的免疫保护作用.     

Insertion of triangulated surfaces into a meccano tetrahedral discretization by means of mesh refinement and optimization procedures

In this paper, we present a new method for inserting several triangulated surfaces into an existing tetrahedral mesh generated by the meccano method. The result is a conformal mesh where each inserted surface is approximated by a set of faces of the final tetrahedral mesh. First, the tetrahedral mesh is refined around the inserted surfaces to capture their geometric features. Second, each immersed surface is approximated by a set of faces from the tetrahedral mesh. Third, following a novel approach, the nodes of the approximated surfaces are mapped to the corresponding immersed surface. Fourth, we untangle and smooth the mesh by optimizing a regularized shape distortion measure for tetrahedral elements in which we move all the nodes of the mesh, restricting the movement of the edge and surface nodes along the corresponding entity they belong to. The refining process allows approximating the immersed surface for any initial meccano tetrahedral mesh. Moreover, the proposed projection method avoids computational expensive geometric projections. Finally, the applied simultaneous untangling and smoothing process delivers a high‐quality mesh and ensures that the immersed surfaces are interpolated. Several examples are presented to assess the properties of the proposed method.     

Certification Institutions Expected to Update and Transfer Information on Present Standards

Standards are the basis for production enterprises to organize production, ex-factory inspection, trade (delivery) and technical exchanges, product certification, quality arbitration and supervision.……     

Flow calorimeter for enthalpy increment measurements on condensed gases

A flow calorimeter for enthalpy increment measurements on condensed gases is presented. A better knowledge of the properties of the liquefied natural gas is needed, and therefore a liquid loop has been designed for our flow calorimeter. The fluid loop in the calorimeter is designed in order to avoid the two-phase region, since two phases would give compositional disturbances in the measurements. The avoidance of the two-phase region is made possible by increasing the pressure of the test fluid after the measurement section, then heating the fluid at super-critical pressure past the critical point. Finally, the fluid is throttled to the low-pressure gas state at the inlet condition of the compressor that circulates the fluid. To perform the pressure increase, a new cryogenic pump has been designed. To evaluate the new equipment, measurements were taken on liquid ethane over the temperature range 146–256 K at pressure between 0.9 and 5.1 MPa.     

Refrigerators Enter a New Era of Health Preservation Frestech initiates the establishment of China's first standard for household sterilization refrigerators

On November 30, 2007, the China Association for Standardization (CAS) held a press conference at Beijing Diaoyutai State Guest House. Leaders from the China Household Electric Appliance Research Institute, the China Household Electric Appliance Association, and the China Consumers' Association attended and made speeches.……     

Optimum-Simulation von Ernteprozessen

Zusammenfassung Bei der Gestaltung von Ernteprozessen — beispielsweise des Prozesses Mähdrusch, Körnertransport und -abnahme — als transportverbundene Fließarbeitsverfahren mit mindestens zwei verschiedenen Arbeitsmitteln gibt es einen großen Optimierungsspielraum. Solche Prozesse sind mehrstufige Bedienungsprozesse mit gemischter Anordnung der Bedienungseinrichtungen. Außerdem besitzen sie stark ausgeprägte räumliche Aspekte. Es wird vor allem gezeigt, welche Merkmale solcher Prozesse in Simulationsmodellen abgebildet sein müssen, welche Zielgrößen in Abhängigkeit von welchen Einflußgrößen unter Berücksichtigung welcher Prozeßbedingungen ermittelt werden sollten und wie der Bewertungsprozeß zur Ermittlung der gewünschten Informationen gestaltet werden muß.     

A study of centrifugal cast high boron high speed steel

In the present study a high‐boron high speed steel (HSS) roll material was designed. Many expensive alloy elements have been substituted by cheap boron alloy, and high‐boron high speed steel roll has been manufactured by centrifugal casting method. The microstructures, mechanical properties and wear resistance of centrifugal casting high‐boron high speed steel roll have been investigated by optical microscopy (OM), scanning electron microscopy (SEM), and X‐ray diffraction (XRD) analysis, hardness test, impact test and wear test. The results indicated that the solidification microstructures of high‐boron high speed steel roll consisted of M₂(B,C), (W,Mo)₂(B,C), M₃(B,C), M₂₃(B,C)₆ type borocarbides and martensite, a small amount of retained austenite. Borocarbides were continuously distributed over the grain boundary. After quenching from 1050 °C, local broken network appeared in partial borocarbides, and fine secondary borocarbide precipitated from the matrix. After tempering from 525 °C, the amount of precipitated borocarbide increased significantly. After heat treatment, the hardness of high‐boron high speed steel roll excelled 60 HRC, and its impact toughness excelled 8.0 J/cm². The single groove steel rolling amount of high‐boron high speed steel rolls increases by 500% than that of bainite cast iron roll, when the rolls are used in K₁ mill housing of bar mill.     

Thixotropy of Systems with Shear Thinning and Plastic Flow Behavior

The definition of the thixotropy is a decrease in viscosity with time in shear and a subsequent recovery of viscosity after the shear deformation is removed.We ...     

Brainstorming for ISO Strategy 2016-2020

正Responding to ISO’s appeal for survey for the to-be-developed ISO Strategy Plan for 2016-2020,SAC hosted a special workshop over the needs and comments of various fi elds on June 8 in Beijing.The workshop was attended by ISO President-elect and Ansteel General Manager Zhang Xiaogang,SAC Vice-Administrator Yu Xinli,and 30 experts from     

CNAS Passes APLAC's PTP Review

正The review group of Asia Pacific Laboratory Accreditation Cooperation(APLAC)carried out a 4-day peer review of the proficiency testing provider(PTP)for China National Accreditation Service for Conformity Assessment(CNAS)from July 28 to 31,2014.CNAS passed the PTP review with excellent performance.Two senior peer reviewers of APLAC made a comprehensive,rigorous and detailed review of CNAS’s PTP accreditation activities,and made the on-spot inspection of the review groups in Shanghai and Shenzhen respectively.