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Y Co-loaded graphitable carbon hollow spheres as anode materials for lithium-ion battery 2008(2)232.Wu Z S;Ren W;Wen L Graphene anchored with Co3O4 nanoparticles as anode of lithium ion batteries with enhanced reversible capacity and cyclic performance 2010(6)引用本文格式：罗飞.褚赓.黄杰.孙洋.李泓.LUO Fei.CHU Geng.HUANG Jie.SUN Yang.LI Hong锂离子电池基础科学问题(Ⅷ)——负。

abbreviated or abridged applicat i on简略申请abnormal karyology异常核型absorbed moisture 吸附水acceptable daily intake可接受的日摄入量acceptable test加速试验acceptance criteria认可标准accuracy准确性accelerated/stress stability studies加速/强力破坏稳定性研究acetylation乙酰化作用achiral assay非手性测定achlorhydric elderly老年性胃酸缺乏症action limits内控限值active components/compound/moiety活性成分active ingredient活性组分adaption to specific culture conditions特定培养条件的适应additives添加剂adjuvant佐剂adventitious agents外源性因子adventitious contaminants 外来污染物adventitious viral or mycoplasma contamination外源性病毒或支原体污染adventitious viruses外源病毒adverse reaction不良反应aerobic microorganisms需氧微生物affinity 亲和力affinity chromatography亲和层析affinity column亲和柱agar and broth琼脂和肉汤aggregates聚合体aggregation聚集allergenic/allergic extracts过敏源抽提物altered conjugated forms改变的结合物形式ambient condition自然条件amino acid composition氨基酸组成amino acid sequence氨基酸序列amino acids氨基酸amino sugars氨基糖amino-terminal amino acids 氨基端氨基酸ammonia production Rates产氨率analyte被测物analytical procedure分析方法animal cell lines动物细胞系animal tissues or organs动物组织或器官anternnary profile触角形状antibiotic resistance genes抗生素耐药基因antibiotics抗生素antibody抗体antibody production tests抗体产生实验antisera抗血清applicant申报者ascites腹水assay含量测定assay procedure定量方法avian鸟类avidity亲和性background背景bacteria细菌batches批次batch-to-batch逐批between-assay variation试验间变异binary fission双数分裂binding assays结合试验bioburden生长量/生物负荷biochemical methods生化方法bioequivalency生物等效性biohazard information生物有害信息biological ativity生物活性biological products生物制品bioreactor生物反应器biotechnological/biological products生物技术/生物产品biotechnological products生物技术产品biphasic curve双相曲线blood plasma fators血浆因子body fluids体液bovine牛bovine spongiform encephalopathy(BSE)疯牛病bracketing括号法breeding conditions饲养条件by-prducts副产物calibrate标化canine犬cap liner瓶帽内垫capillary electrophoresis毛细管电泳carbohydrate碳水化合物carboxy-terminal amino acids碳基端氨基酸carrier载体/担体catalysts催化剂cell bank细胞库cell bank system细胞库系统cell banking procedures细胞建库过程cell banking system细胞库系统cell culture-derived impurities来源于细胞培养基的杂质cell cultures细胞培养物cell expansion细胞扩增cell lines细胞系cell metabolites细胞代谢物cell pooling细胞混合cell substrate-derived impurities来源于细胞培养基质的杂质cell substrates细胞基质cell viability细胞活力cell–derived biological products细胞来源的生物制品cell fusion细胞融合cellular blood components血细胞成分cemadsorbing viruses红细胞吸附病毒characterization and testing of cell banks细胞库鉴定及检测charcoal活性炭charge电荷chemical actinometric system化学光化线强度系统chemical reactivity化学反应性chemical syntheses化学合成chemically inert化学惰性chewable tablets咀嚼片chiral impurities手性杂质chromatograms色谱图chromatographic behavior色谱行为chromatographic procedures色谱方法chromatography columns色谱柱circular dichroism圆二色性clearance studies清除研究climatic zones气候带clinical research临床研究clinical trial application临床实验申请cloning克隆closure闭塞物code number编号coding sequence编码序列coefficient of variance变异系数collaborative studies协作实验研究colony isolation菌落分离colony-stimulating factors集落刺激因子combination product复方制剂components成分confidence interval置信区间confidence limits 可信限confirmatory studies确认研究conformance to specifications符合规范conformation构型conjugat ed product连接产物consistency一致性container容器container/closure容器/闭塞物container/closure integrity testing 容器/密封完整性实验contaminants污染物content uniformity含量均匀性control methodology控制方法学controlled released product控制制剂conventional vaccines传统疫苗conventional live virus vaccines传统的活病毒疫苗cool white fluorescent冷白荧光灯correction factor校正因子correlation coefficient相关系数covalent or noncovalent共价或非共价creams霜剂cross-contamination交叉污染cross-reactivity交叉反应cryopreservation冷冻保存cryoprotectants防冻剂crystals晶体culture components培养基成分culture media/medium培养基cyanogens bromide溴化氰cytogenetic细胞遗传学的cytokines细胞因子cytopathic细胞病的cytoplasmic A-and R-type particles细胞浆a型和r型颗粒dark control暗度对照deamidation 去氨基deaminated去酰胺化的decision flow chart/tree判断图definable and measurable biological activity明确和可测定的生物学活性degradant降解产物degradation降解degradation pathway降解途径degradation product降解产物degradation profile降解概况degree of aggregation 凝集程度degree of scatter离散程度delayed –release延迟释放deleterious有害的delivery systems给药体系derivatives衍生物description性状detection limit检测限度dilivery systems释放系统dilution ratio稀释倍数dimmers二聚体diode array二极管阵列diploid cells二倍体细胞dissociation解离dissolution testing 溶出实验dissolution time溶出时间dosage form剂型downstream purification下游纯化drug product制剂drug product components制剂组方drug substances原料药ectromelia virus脱脚病病毒elastomeric closures橡皮塞electron microscopy (EM)电镜electrophoresis电泳electrophoretic pattern电泳图谱elution profile洗脱方案embryonated eggs鸡胚enantiomer对映体enantiomeric对映异构体enantioselective对映体选择性encephalomyocarditis virus(EMC)脑心肌炎病毒endogenous agents内源性因子endogenous retrovirus内源性逆转录病毒endotoxins内毒素end-product sterility testing 最终产品的无菌试验enhancers增强子enveloped RNA viruses包膜RNA病毒environmental factors环境因素enzymatic reaction rates酶反应速率enzyme酶epitope表位Epstein-Barr virus(EBV)EB病毒equine马Erythropoietins促红细胞生成素ethnic origin种族起源eukaryotic cell真核细胞ex vivo体外excipient赋形剂excipient specifications赋形剂规范expiration date/dating 失效日期exposure level暴露程度exposure period光照时间expression constract表达构建体expression system表达系统expression vector表达载体extended-release延时释放extent of the virus test病毒测试程度extinction coefficient消光系数extrachromosomal染色体外extraneous contaminants 外源性污染物exprapolation外推法fermentation发酵fermantation products发酵产物fill volume装量filter aids过滤介质final manufacturing最终生产finished product成品flanking region侧翼区forced degradation testing 强制降解物质foreign matter异质性物质formal labeling 正式标签formal stability studies正式的稳定性研究formulation处方/配方fragmentation片段化freeze-dried product冻干产品friability脆硬度fungi真菌fusion partnters融合伴侣fusion protein融合蛋白gel filtration凝胶过滤gene amplification基因扩增gene therapy基因疗法generation of the cell substance细胞基质的产生genetic manipulation基因操作genomic dinucleotide repeats基因组双核苷酸重复数genomic DNA基因组DNAgenomic polymorphism pattern基因组形态类型glucose consumption rates耗糖率glycoforms糖化形式glycosylation糖基化goegrapgical origin地理起源growth factors生长因子growth hormones生长激素guanidine胍hamster antibody production (HAP) test仓鼠抗体产生实验Hantaan virus汗坦病毒Hardness硬度heavy metals重金属heparins肝素herbal products 草药herpes virus疱疹病毒heterogeneities异质性heterohybrid cell lines异种杂交细胞系high-resolution chromatography高分辨色谱homogeneity均一性host cell宿主细胞host cell banks宿主细胞库host cell DNA宿主细胞DNAhost cell proteins宿主细胞蛋白质hot-stage microscopy热阶显微镜human diploid fibroblasts人二倍体呈纤维细胞human polio virus人脊髓灰质炎病毒human tropism人向性humidity湿度humidity-protecting containers防湿容器hybridization techniques杂交技术hybridoma cell杂交瘤hybridomas水解物hydrolysates水解物hydrolytic enzymes水解酶hydrophobicity疏水性hygroscopic吸湿性idetification/identity鉴别immediate container/closure直接接触的容器/密闭物immediate pack内包装immediate release立即释放immortalization激活immune spleen cells免疫脾细胞immumoassay免疫检测immunochemical methods免疫化学方法immunochemical properties免疫化学性质immunoelectrophoresis免疫电泳immunogenicity免疫原性immunological interations免疫相互作用immunoreactivity免疫反应性impurity profile杂质概况in vitro and in vivo inoculation tests体内和体外接种试验in vitro assay体外检测in vitro cell age体外细胞传代期in vitro lifespan体外生命周期in vitro tests体外试验in vivo 体内in vivo assays体内检测inactivated vaccine灭活疫苗indentification test鉴别试验indicator cell指示细胞indicator organisms指示菌indoor indirect daylight室内间接日光inducers诱导剂infectious agents感染性因子influenza virus流感病毒inhalation dosage forms吸入剂型in-house内部的in-house criteria内控标准in-house primary reference material内部一级参比物质in-house working reference material内部参比物质initial filing原始文件initial submission最初申报initial text最初文件inoculation接种inorganic impurities无机杂质inorganic mineral无机矿物质inorganic salts无机盐in-process acceptance criteia生产过程认可标准in-process controls生产过程中控制in-process testing生产过程中检测insect昆虫insulins胰岛素intake摄入intended effect预期效果intended storage period预期的储藏期intentional degradation人为降解interactions相互作用interferon干扰素interleukins白细胞介素intermediate中间体intermediate precision中间精密度intermediates半成品international reference standards国际参比标准品intra-assay precision间隙含量精密度intracytoplasmic细胞浆内introduction of virus病毒介入inverted or horizontal position倒立或水平位置ion-exchange离子交换ionic content离子含量isoelectric focusing/isoelectrofocusing等电聚焦isoenzyme analysis同工酶分析isoform pattern异构体类型isomerized异构化的Jp /Ph.Eur./Usp.日本药局方/欧洲药典/美国药典K virus K病毒Karyology胞核学laboratory scale实验室规模lactate production rates乳糖产生速率litric deheydrogenase virus(LDM)乳酸脱氢酶病毒leachables沥出物ligand配位体/配体light光照light resistant packaging避光包装limit for in vitro cell age细胞体外传代限度limit of acceptance可接受的限度limit of in vitro cell age体外细胞代次limit test限度试验limulus amoebocyte lysate鲎试剂linear relation ship线性关系linearity线性liquid nitrogen液氮liquid oral dosage forms液体口服制剂live vaccine活疫苗living cells活细胞logarithmic scale对数级long term test长期试验long-time and accelerated stability长期和加速稳定性试验losses of activity活性丧失lot release批签发low molecular weight substances低分子量物质lower-observed effect level(LOEL)能观察到反应的最低量lymphocytic choriomeningitis virus(LCM)淋巴细胞性脉络丛脑膜炎病毒lyophilised cakes冻干粉饼lysate of cells细胞溶解物mammalian哺乳类manufacturing scale生产规模marker chromosome标志染色体marketing pack上市包装mass重量mass balance质量平衡mass spectrometry质谱master cell bank (MCB)主细胞库material balance物质平衡matrix基质、矩阵matrix system矩阵法matrixing 矩阵化设计maximum daily dose每日最大剂量mean kinetic temperature平均动力学温度metazoan cell culture后生动物细胞培养microbial cell culture微生物细胞培养microbial cells微生物细胞microbial contamination微生物污染microbial expression system微生物表达系统microbial limits微生物限度microbial metabolites微生物代谢物microbial proteases微生物蛋白酶microbial vaccine antigens微生物疫苗抗原microbiological testing微生物学试验minimum exposure time最低作用时间minimum of pilot plant试验规模minute virus of mice小鼠小病毒mirror image镜像mismatched S-S linked错连的S-S键mork run空白对照试验modified-/modifying release修饰释放modifying factor修正因子moisture level水分molar asorptivity克分子吸收molecular characteristics分子特性molecular confirmation分子构型molecular entities/entity分子实体molecular size分子大小monoclonal antibody单克隆抗体morphological analysis形态学分析mouse antibody production (MAP) test小鼠抗体产生试验mouse cytomegalovirus(MCMV)小鼠巨细胞病毒mouse encephalomyelitis virus(GDVⅡ)小鼠脑脊髓炎病毒mouse hepatitis virus(MHV)小鼠肝炎病毒mouse rotavirus(EDIM)小鼠小轮状病毒MuLV murine leukemia virus鼠白血病病毒murine hybridoma cell lines鼠杂交瘤细胞系mutations突变mycoplasma支原体myeloma cell line骨髓瘤细胞系national or international reference material国家或国际参比物质near ultraviolet lamp近紫外灯neural sugars中性糖new chemical entry 新化学体new dosage form新剂型new drug products/produce新药制剂new drug substance新原料药new molecular entities新分子体no effect level不产生反应的量noncovalent/convalent force非共价/共价键non-enveloped viruses非包膜病毒non-mammalian animal cell lines非哺乳动物细胞系non-recombinant cell-culture expression systems非重组细胞培养表达系统non-recombinant products/vaccines非重组制品/疫苗non-specific model virus 非特异模型病毒no-observed effect level不能观察到反应的量N-terminal sequencing N-端测序nuclear magnetic resonance核磁共振official procedure正式方法ointments软膏oligosaccharide pattern低聚核苷酸opacity浊度origins of replication复制起点osmolality摩尔渗透压浓度outdoor daylight室外阳光oxidation氧化oxygen consumption rates耗氧率package包装parainfluenza virus副流感病毒parallel control assays平行对照分析parent stability Guideline稳定性试验总指导原则patrental cell line母细胞系parenterals非肠道制剂particle size粒度particulate matter微粒parvoviruses细小病毒passage history of the cell line细胞系的传代史pathogenic agents致病因子pathogenicity致病性patterns of degradation降解方式peptide肽peptide map肽图percent recovery回收率periodic/skip testing定期检验/抽验permitted daily exposure允许的日接触量phage typing 噬菌体分型pharcodynamic studies药效学研究pharmacopoeial药典pharmacopoeial specifications药典规范pharmacopoeial standards药典标准phenotypic表型phosphorylation磷酸化作用photostability testing光稳定性试验physicochemical changes理化改变physicochemical methods物理化学方法physico-chemical properties物理化学特性pilot-plant scale试生产规模/中试规模piston release force活塞释放力piston travel force活塞移动力pivotal stability studies关键的稳定性研究plaque assays菌斑测定plasmid质粒plasmid banks质粒库plasminogen activators纤溶酶原激活素pneumonia virus of mice小鼠肺炎病毒Poisson distibution泊松分布polymorphic form多克隆抗体polymorphse chain reaction（PCR）聚合酶链式反应polymorphic form多晶性型polymorphs多晶型polyoma virus多瘤病毒pooled harvest集中回收population doubling细胞鼠倍增/群体倍增porcine猪post-approval批准后post-translational modification翻译后修饰post-translationally modified forms翻译后修饰形式potency效价potent功效potential adverse consequences潜在的不良后果potential excipients准赋形剂potential impurity潜在杂质potential new drug products准新药制剂potential new drug substances准新原料药potentiometric titrimetry电位滴定powders粉剂power outages and human error断电和人为错误preamble引言pre-approval or pre-liscense stage批准前或发证前阶段precision精密度preclinical and clinical studies临床前和临床研究precursors前体preliminary assessment初步评估preliminary cell bank初级细胞库preparation制剂preservative防腐剂primary cells原代细胞primary stability data主要稳定性数据primary stability study/formal study/formal stability study主要稳定性研究/正式研究/正式稳定性研究primary structure一级结构primer引物priming regimen(s)接种方式probability概率process characterization studies工艺鉴定研究process controls工艺控制process optimization工艺优化process parameters工艺参数process validation工艺确证process –related impurities工艺相关杂质product-related impurities产品相关杂质progenitor祖细胞prokaryotic cell原核细胞promoters启动子proposed commercial process模拟上市protected samples避光样品proteins蛋白质分析技术proteolysis蛋白水解protocol方案pseudopolymorphs伪多晶体pseudorabies virus假狂犬病毒purification纯化purified antigens纯化抗原purity纯度purity test纯度试验pyrogens热原试验qualification界定qualified 合格的quality standards质量标准quantal methods质反应测定法quantitation limit定量限度quantitative characteristics定量参数quantitative detection定量检测quantitative infectivity assays感染性定量测定quantitative method定量方法quantitative test定量试验quantitative virus病毒定量分析quantity含量racemate消旋体radiometers/lux meters测光仪/照度仪radiopharmaceutical放射性药物range范围rat antibody production (RAP) test大鼠抗体产生试验rationale基本原理raw material原材料raw material testing原材料测试rDNA technology重组DNA技术rDNA-modified cell substrates重组DNA修饰的细胞基质reagent试剂、反应物real condition真实条件real time 真实时间rebank 再建库receptor受体reclone再克隆recombinant cell-culture expression systems 重组细胞培养表达系统recombinant DNA protein products重组DNA蛋白质产品recomibinant-DNA-derived product重组DNA制品recombinant protein重组蛋白质reconstitution重新溶解redispersibility再分散性reduction factors下降因子reference material参比物质reference standard参比标准品regimen方案registration application注册申请regression analysis回归分析regulator/regulatory agencies管理机构related substances有关物质release limit出厂限度“relevant” viruses and “model” viruses“相关”病毒和“模型”病毒reovirus type3(Reo 3)呼吸肠病毒repeatability重复性reproducibility 重现性residual solvents残留溶剂residual sum of squares溶剂残留量resolution test分离度试验response factor响应因子restriction endonuclease mapping限制性内切酶图谱restriction fragment length polymorphism限制性片段长度多态性resuspension再悬浮retention time保留时间retest date再试验日期reverse transcriptase(RT)反转录酶reversed-phase chromatography反相色谱reverse-phase liquid chromatography反相液相色谱revived cells复苏的细胞rheological properties流体学特性risk-benefit analysis利弊分析robustness耐用性rodent retrovirus啮齿类动物逆转录病毒sampling采样scale-up放大scaling down缩小规模scope范围scrapie瘙痒病screening tests筛选试验SDS-PAGE/SDS-polyacrylamide gel electrophoresis十二烷基硫酸钠-聚丙烯酰胺凝胶电泳sealed ampoules密封安瓿secondary structure二级结构self-replicating agents自我复制因子semi-synthetic products半合成产品Sendai virus仙台病毒Sensitivity灵敏度Senescence老化Separation分离Serum血清Shear切变shelf life货价寿命shipment运输sialic acids唾液酸signal-to-noise信噪比Sindbis virus新德比病毒single-dose and multiple-dose packages单剂量和多剂量包装single-point measurements单点测定single-tiered banking system单级细胞库系统size exclusion chromatography分子排阻色谱skip lot testing随机试验slope of the regression line回归线的斜率solid oral doseage固体口服制剂solvates溶剂化物solvation溶剂化作用solvent溶剂species物种specific gravity比重specific objectionable bacteria控制菌specification规范specification limit规范限度specification –check质控规范specification-release出厂规范specifications规范specificity专属性specified impurities特定杂质specified light exposure特定的光照spectroscopic profiles光谱图spiked samples加料样品spiking experiments叠加试验splicing sequences剪接序列stabilizers稳定剂stability data稳定性资料stability evaluation稳定性评价stability proticol稳定性方案stability study duration稳定性试验期限stability testing稳定性试验stability-indicating profile反应稳定性指标standard deviation标准差standard stock solution标准储备液starting materials起始物statement/labeling说明/标签statistical analysis统计学分析sterility无菌storage condition放置条件strains品系stress condition强力破坏试验条件stress testing强力破坏试验storage conditions储存条件structural heterogeneity结构异质性subcultivations传代培养sulfhydryl groups and disulfide bridges巯基和二硫键sulfoxidation硫酰化support information辅助性资料surrogate test替代试验surrogates替代物suspensions混悬剂swine猪synthesis合成synthetic peptides合成肽syringeability灌注功能systemic exposure全身接触tablet cores片芯tandem repeats串联重复target molecule靶分子temperature changes温度变化terminology术语tertiary structure三级结构test criteria试验标准test intervals试验间隔test parameters试验参数testing frequency试验次数texture质地the method of least squares最小二乘法threshold limits阈值thymic virus胸腺病毒tip cap removal force滴帽移动力tissue-culture-infectious-dose(TCID)组织培养感染剂量titration滴定法tolerable daily intake可耐受的日摄入量toolan virus(Hi)图兰病毒topical formulations局部用药处方toxic impurity毒性杂质toxin毒素tranfection of matazoan cells后生动物细胞的转染transcrption转录transdermal systems透皮吸收系统transfection转染transfomation转化translational fidelity翻译的忠实性transmission electron microscopy电透镜transparent cover透明盖子truncated forms截短形式tumor necrosis factor肿瘤坏死因子tumorigenicity致瘤性two-tiered cell bank两级细胞库uncloned cell population未克隆的细胞群unicellular life forms单细胞生命形式unidentified impurities未确定杂质uniformity of content 含量均匀度uniformity of dosage units剂量单位的均匀度uniformity of fill装填均匀度uniformity of mass质量均匀度unitage单位universal tests/criteria常规试验/标准untransfected recipient cell line未转染的受体细胞系UV/visible wavelength紫外可见光波长Vacinnes疫苗Validation论证variant sequences变异序列variants 变异体vector载体vehicle载体/溶酶vesicular stomatitis virus小囊状口腔炎病毒viral clearance病毒清除viral clearance studies病毒清除研究viral contamination病毒污染viral geneme病毒基因组viral infectivity病毒感染性viral safety evaluation病毒安全评估viral vicinnes病毒性疫苗virucidal buffers杀病毒缓冲液virus load 病毒浓度virus titer病毒滴度viscosity粘度visible particulates可见颗粒visual appearance外观visual evaluation直观评价vitamins 维生素water of hydration结晶水well-defined testing program确定的试验项目western blot免疫印迹whole blood全血within-assay variation试验内变异working cell bank工作细胞库yeast酵母y-intercept y轴上的截距。

[作者简介]田忠山,男,内蒙古工业大学外国语学院助教,在读硕士研究生。

The Approach of Contra st i n Character i zati on of H a m let○Tian Zhongshan(Foreign L anguage Institu te,InnerM ongolia Polytechn ic U niversity,Hohhot,InnerM ongolia,Ch ina 010010)　[Abstract]　This paper is a study of the app r oach of contrast,which is e mbodied in Shakes peare’s master p iece,Ham let .By the comparis on bet w een Ha m let’s p r of ound thought and Laertes’s rashness and Claudius’s wickedness and oldHa m let’s l oftiness,we can see that contrast is one of the maj or app r oaches adop ted by Shakes peare in this work .This is an effective way t o make the characters’distinguishing pers onalities become more distinguishable .W hile Shakes peare’s great 2ness doesn’t lie in the adop ti on of this common way of writing,it lies in his flexibleness and masteries in his writing in the p r ocess of e mp l oying the app r oach of contrast .　[Key words]　contrast;characterizati on;Ham let　[中图分类号]I 10614 [文献标识码]A [文章编号]167228610(2007)1220072203 Shakes peare,the most outstanding dra matist all o 2ver the world,is very skilled in creating full bl ooded,rep resentative and vivid characters .The characters un 2der his pen see m t o be gifted with lives in his p lays .Among his 37dra mas,Ham let is one of the most re 2markable,in which Shakes peare shows his great talent of characterizati on .The characters in this p lay are all very i m p ressive .Each of the m has their distinctive in 2dividuality,f or exa mp le,the melancholy of Ha m let,the weakness and innocence of Ophelia,the wicked 2ness of Claudius,the pedantry of Pol onius,and s o on .Even Rosencrantz and Guildenstern become world 2wide household na mes,which stand for disl oyalty and treachery .T o achieve this vividness in dep icting characters,Shakes peare adop ts diversified skills,a mong which contrast is a very cons p icuous one .I n literary works,the e mp l oying of this app r oach can leave a very shar p and vivid i m p ressi on by menti oning t w o co mp letely dif 2ferent pers onages in one work .This paper is just goingt o discuss the app r oach of contrast in characterizati on e mp l oyed by Shakes peare in Ham let .Among the gallery of characters portrayed by Shakes peare in Ham let ,the most notable one is Ha m let unquesti onably .Shakes peare shows with great enthusi 2as m his p r of ound thought,his res olute deter m inati on and his l ofty s oul .Those distinguishing features be 2come more distinguishable in contrast against other characters’pers onalities .I n this p lay,several p l ots are woven t ogether .Both Ha m let and Laertes undertake t o avenge their fa m ilies on killing,whereas,they convey comp letely different pers onalities in their acti ons .I n the p r ocess of Ha m let’s revenge,his delay in acti on just shows his outstanding intelligence .A t the beginning of the st ory,he dis p lays his s orr ow f or his father ’s death and his mother’s hasty marriage .A t the sa me ti m e,in his deep s oul,he sus pects the real cause of his father’s death .Because he is s o intelligent he can see thr ough things .That’s why when the ghost tells hi m :27L I TERAT URE Tian Zhongshan /The App r oach of Contrast in Characterizati on of Ham letThe ser pent that sting they father’s lifeNow wears his cr own.(Shakes peare.V.38239) his res ponse is:“Oh,my p r ophetic s oul.”(V.60).I n s p ite of this str ong intuitive sus p ici on,he doesn’t ex2 hibit it.He doesn’t rush int o the cha mber of the as2 sumed murderer’s with a cr owd of mobs and questi ons the king directly:“Give me my father!”(,5),just as what Laertes does.Considerable grief at his father’s death and gr oundless g ossi p make Laertes l ose all his reas ons and s oberness,and he is driven nearly mad by the str ong desire of revenge:T o this point I stand,That both worlds give t o negligence,Let come what comes,only I’ll be revengedMost thor oughly f or my father.(..1332136)These words are the best portrayal of Laertes’s rashness;everything he does is urged by passi on and i m pulsiveness.W hile under the sa me circumHa m let’s perf or mance is sur p risingly composed.W hen the ghost comes and reveals hi m the murder of his fa2 ther,he is over whel m ed by shock and indignati on,but he is not conquered by these e moti ons.H is p rudent nature drives hi m t o questi on what others take f or gran2 ted,he says:The s p irit that I have seenMay be the devil,and the devil hath powerT’assu me a p leasing shape;(..5852587)So he has a p lay acted which rese mbles the death of old Ha m let,in order t o justify the king’s f oul deeds and find convincing evidence for the unnatural murder. W hen the guilty Claudius stands up and leaves the hall in fear bef ore the p lay ends up,everything becomes clear as clear can be.However,Ha m let doesn’t take i m mediate acti on when a good opportunity comes. Claudius,who is stricken by guilty consci ous is knee2 ling and atte mp ting t o p ray,al one.He could have kill2 ed hi m on the s pot,but when he dra ws his s word,he hesitates:Now m ight I do it pat,now he is a2p raying;And now I’ll do it,and s o he goes t o heaven,And s o I a m revenged.That would be scanned.(..71274)He gives up this extre mely favorable opportunity, because he wants t o destr oy his ene my’s s oul as well as body.He al w ays thinks the consequence of his deeds over and over again,that leads t o his delay in acti on, which just reflects his p r ofound pared with Ha m let,Laertes appears s o si m p le2m inded and reck2 less,and these weaknesses are taken advantages of by the crafty,sha meless king.Consequently,Laertes be2 comes the access ory t o the murder of p rince Ha m let.Fr om above,it is easy t o conclude that Ha m let’s intelligence and p rudence stand more vivid in contrast against Laertes’s rashness and naivety.The comparis on bet w een the t w o’s pers onalities creates a shar p contrast. Not only that,exa m ining the dra ma on the whole,it can be f ound that the app r oach of contrast p lays really an i m portant r ole in the portrayal of the i m age of Ha m2 paring with the weakness and purity of Ophel2 ia his res oluti on and strength appear more striking;in comparis on with the wicked Claudius,Ha m let’s i m age see m s even l oftier and nobler.T o s peak of Claudius,the e mp l oying of contrast is more obvi ous in the p r ocess of the creating of this i m2 age.It is known t o all,in this p lay Claudius pers oni2 fies all the f orces of evil.He pois ons his own br other t o death,and then marries his sister2in2la w unscrupul ous2 ly.H is ugliness see m s more p r ojecting in contrast with old Ha m let’s l oftiness.This f oll owing sentence appears in an i m passi oned s olil oquy of Ha m let:So excellent a king that was t o this,Hy peri on t o a Satyr.(..1392140)I n this s olil oquy he exhibits his extreme disgusting t o his mother’s marriage and shows his str ong distain t o his uncle.No more sentences in this seg ment can be found t o describe how ugly and repulsive Claudius is. The wonderfulness of e mp l oying contrast is it leaves a s pace f or the readers t o i m agine:how different old Ha m let is fr om Claudius is just as how different Hype2 ri on is fr om Satyr.A s the st ory g oes on,Ha m let’s a2 bom inati on t oward his uncle gr ows even str onger,when his cri m e has been justified.I n the queen’s cha mber,37语文学刊(高教・外文版) 2007年第12期Ha m let reveals Claudius’s baseness t o her which ren2 ders the queen’s heart br oken:Look here,upon this p icture,and on thisA combinati on and a f or m indeedW here every god did see m t o set his seal t o give the world assurance of a manHere is your husband,like m ild ear(..53256)By this shar p contrast any more descri p ti on is su2 perfluous,and the i m age of a des p icable king vividly appears in fr ont of the readers.By the analysis of the t w o i m ages—Ha m let and Claudius,the clear conclusi on is that the app r oach of contrast is one of the maj or app r oaches e mp l oyed by Shakes peare in the p r ocess of characterizati on in Ham2 let.Nevertheless,contrast is just a very common ap2 p r oach of characterizati on,which can be found in nu2 mer ous literary works in any era,but in Shakes peare’s hand,it becomes a most flexible and effective method. W hile what we shouldn’t ignore is that contrast is just one of the app r oaches of characterizati on adop ted byShakes peare,there are still other ways such as p sycho2 l ogical analysis.So it is not inapp r op riate t o say that Shakes peare is a great master of characterizati on.Under Shakes peare’s hands,even the most insignificant fig2 ures are fa m iliar t o the readers,and he hi m self is con2 sidered as the greatest dra matist in the world.【References】[1]Guo Qunying.B ritish L iterature[M].Foreign LanguageTeaching Research Press,2001.[2]L iu B ingshan.A Short H istory of English L iterature[M].He Nan Peop le’s Publishing House,1973.[3]Luo J ingguo.A N e w A nthology of English L iterature[M].Bei J ing University’s Publishing House,1996.[4]Shakes peare,W illia m.Ham let[M].Eds Cedric W alts,Crib street,W are,Hertfordshire:Words worth Editi onsL i m ited Cu mberland House,1992.[5]W u W eiren.H istory A nd A nthology of English L iterature[M].Foreign Language Teaching and Research Press,2000(revised ed.)对照手法在莎士比亚的代表作《哈姆雷特》中的运用田忠山(内蒙古工业大学外国语学院,内蒙古呼和浩特010010)[摘　要]　通过对哈姆雷特思想的深邃与雷欧提斯的鲁莽、哈姆雷特的高贵与克罗狄斯的卑鄙的对比,可以得出这样的结论:对照手法是莎士比亚在这部作品中采用的主要手法。

计算机专业术语中英⽂对照计算机专业术语对照Aabstraction layer，抽象层access，获取，存取acoustic coupler，声⾳耦合器Active Directory，活动⽬录Acyclic Dependencies Principle，⾮循环依赖原则（ADP）acyclic digraph，有向⽆环图Adaptive Code，⾃适应代码Add Parameter，添加参数ADSL，Asymmetrical Dingital Subscriber Loop，⾮对称数字⽤户环线affinity，绑定affinity group，地缘组agent，代理agent-based interface，代理⼈界⾯Agile，敏捷⽅法论agile practice，敏捷实践agile peocess，敏捷流程agility，敏捷性AI，Artificial Intelligence，⼈⼯智能air waves，⽆线电波algorithm，算法analog，模拟的animation，动画annotation，注解，注释answering machine，电话应答机antenna，天线anti-pattern，反模式APM，异步编程模型（Asynchronous Programming Model）Apocalyptic defect，灾难缺陷application，应⽤，应⽤程序，应⽤软件application life cycle，应⽤程序⽣命周期application pool，应⽤程序池Application Programming Interface，应⽤程序编程接⼝（API）architecture，体系机构，结构architecture decay，架构腐坏Architecture Style，架构风格ARPA，Advanced Research Projects Agency，（美国国防部）⾼级研究计划署ARPAnet，ARPA⽹Arrange-Act-Assert，准备-执⾏-断⾔（AAA）artifact，构建物4ASF，Apache Software Foundation 的简写Aspect-Oriented Programming，⾯向切⾯编程（AOP）aspect ratio，屏幕⾼宽⽐assembly，程序集Asynchronous Programming Model，异步编程模型（APM）ATM，asynchronous transfer mode，异步传输模式atomic opreation，原⼦操作atomic transaction，原⼦事务atomicity，原⼦性attribute，特性augmented reality，增强实现authentication，⾝份验证authorization，授权automated unit testing，⾃动化单元测试automation，⾃动化autonomous，独⽴性availability，可⽤性availability set，可⽤性集AZs，可⽤性区域（Availability Zones，亚马逊 AWS 中数据中⼼的叫法）4BBackend as a Service，后端即服务（BaaS）backpane，底板backward compatibility，向后兼容性bandwidth，带宽bar code，条形码Base Class Library，基类库（BCL）baseline，准线baud，波特BCL，基类库（Base Class Library）bear，熊behavior，⾏为behavior preserving program transformations，⾏为保留式程序转换1 Behavioral error，⾏为错误BFF，为前端服务的后端（Backends For Frontends）4Big Ball of Mud，⼤泥球（BBM）big data，⼤数据Big Design Up Front，⼤优先设计（BDUF）binary，⼆进制的binochlar，双⽬并⽤的bit，⽐特Bit-field，位域bitnik，⽐特族blob，BLOBblock，阻断block blob，块 BLOBBlockchain as a Service，区块链即服务（BaaS）bottleneck，瓶颈bounded context，边界上下⽂、界限上下⽂4box，装箱bps，bits per second，⽐特/秒breakpoint，断点broadcast，（⽆线电或电视）⼴播Broken Hierarchy，⽀离破碎的层次结构2Broken Modularization，拆散的模块化2brownfield project，⾏进中项⽬Browser Object Model，浏览器对象模型（BOM）browser-server，浏览器-服务器bug，缺陷built-in，内置的，内建的；嵌⼊的；内置bulkhead，舱壁4business intelligence，商业智能business layer，业务层business logic layer，业务逻辑层busy (status)，忙（状态）；繁忙（状态）byte，字节Ccable，电缆Cache/Caching，缓存call stack，调⽤堆栈callout box，标注框camelCase，camel ⼤⼩写canary releasing，⾦丝雀发布4carbon copy，复写本，副本；抄送（CC）carriage return，回车Cascading Style Sheets，层叠样式表（CSS）catastrophic failover，灾难性故障转移4CD，持续交付（Continuous Delivery）4CDC，消费者驱动的契约（Customer-Driven Contract）4CDN，内容分发⽹络（Content Delivery Network）cell，单元cellular telephone，移动电话Central Processing Unit，中央处理器（CPU）certificate，（数字）证书Certificate Authority，证书认证机构Change Bidirectional Association to Unidirectional，将双向关联改为单向关联1Change Point，修改点：需要往代码中引⼊修改的点Change Reference to Value，将引⽤对象改为值对象1Change Unidirectional Association to Bidirectional，将单向关联改为双向关联1Change Value to Reference，将值对象改为引⽤对象1channel，信道，频道character，字符Characterization test，特征测试：描述软件某部分的当前⾏为的测试，当你修改代码时能够⽤来保持⾏为check in，签⼊check out，签出chip，芯⽚choreography，协同CI，持续集成（Continuous Integration）4cipher，密码claim，声明class definition，类定义CLI，公共语⾔基础结构（Common Language Infrastructure）client-server，客户端-服务器clone，克隆，复制cloud computing，云计算cloud service，云服务CLR，公共语⾔运⾏时（Common Language Runtime）CLS，公共语⾔规范（Common Language Specification）cluster，集群clustered index，聚集索引CMS，内容管理系统（Content Management System）co-occurring smells，同时出现的坏味2coaxial cable，同轴电缆COBIT，信息和相关技术的控制⽬标，Control Objectives for Information and Related Technology4 CoC，更改开销（Cost of Change）code smell，代码味道Collapse Hierarchy，折叠继承关系1comcurrency，并发command，命令command prompt，命令⾏提⽰Command/Query Responsibility Segregation，命令/查询职责分离（CQRS）Command/Query Separation，命令/查询分离（CQS）commingled bits，混合的⽐特communication，通信community，社区committed，已提交（的）Common Intermediate Language，公共中间语⾔Common Language Infrastructure，公共语⾔基础结构（CLI）Common Language Runtime，公共语⾔运⾏时（CLR）Common Language Specification，公共语⾔规范（CLS）Common Type System，公共类型系统（CTS）common name，通⽤名称compatibility，兼容性Competing Consumer pattern，消费者竞争模式4Component Object Model，组件对象模型（COM）composite formatting，复合格式化Composite Pattern，复合模式concurrency conflicts，并发冲突concurrency mode，并发模式conditional compilation，条件编译conditional compilation statement，条件编译语句configuration，配置，设置connection string，连接字符串Consolidate Conditional Expression，合并条件表达式1Consolidate Duplicate Conditional Fragments，合并重复的条件⽚段1consistenct，⼀致性constructor，构造函数container，容器Container As A Service，容器即服务（CaaS）4content，内容context，上下⽂contextual keyword，上下⽂关键字continuous integration，持续集成contribute，贡献Contributor License Agreement，贡献者许可协议convention，约定covariance，协变contravariance，逆变convert，转换Convert Procedural Design to Objects，将过程化设计转化为对象设计1cookie，Cookiecore，内核；.NET Core 的简写（能且仅能与 .NET Framework 的简写nfx同时出现，作如nfx/core，单独使⽤时应为全称.NET Core）corruption，损毁Cosmetic issue，外观上问题Cost of Change，更改开销（CoC）COTS，现成的商业软件（Commercial Off-The Shelf）4counterpoint，对位4Coupling count，耦合数：当⼀个⽅法被调⽤时传给它以及从它传出来的值的数⽬。

Synthetic natural gas from CO hydrogenation over silicon carbide supported nickel catalystsYue Yu a ,b ,Guo-Qiang Jin a ,Ying-Yong Wang a ,Xiang-Yun Guo a ,⁎a State Key Laboratory of Coal Conversion,Institute of Coal Chemistry,Taiyuan 030001,PR China bGraduate University of the Chinese Academy of Sciences,Beijing 100039,PR Chinaa b s t r a c ta r t i c l e i n f o Article history:Received 19March 2011Received in revised form 23July 2011Accepted 1August 2011Available online xxxx Keywords:CO methanation Ni/SiC catalyst Ni/TiO 2catalyst StabilitySilicon carbide supported nickel catalysts for CO methanation were prepared by impregnation method.The activity of the catalysts was tested in a fixed-bed reactor with a stream of H 2/CO =3without diluent gas.The results show that 15wt.%Ni/SiC catalyst calcined at 550°C exhibits excellent catalytic activity.As compared with 15wt.%Ni/TiO 2catalyst,the Ni/SiC catalyst shows higher activity and stability in the methanation reaction.The characterization results from X-ray diffraction and transmission electron microscopy suggest that no obvious catalyst sintering has occurred in the Ni/SiC catalyst due to the excellent thermal stability and high heat conductivity of SiC.©2011Elsevier B.V.All rights reserved.1.IntroductionNature gas is a clean fuel in fossil fuels and it has a higher calori fic value comparing with petroleum and coal.In recent years,the production of synthetic natural gas from coal and solid dry biomass has been a concern due to the rising price and exhaustion of natural gas [1,2].As one of the most essential steps in the production of SNG [1],the methane synthesis from carbon monoxide and hydrogen has been paid more attention.Different types of methanation catalysts have been developed and widely investigated since the reaction was first reported by Sabatier and Senderens in 1902[3].Although noble metal catalysts show higher activity [4,5],Ni-based catalysts are widely applied due to the low-cost and good availability [1].Many materials such as TiO 2,Al 2O 3,CeO 2,SiO 2,ZrO 2,MgO,YSZ and MgAl 2O 4have been investigated as the support of nickel catalysts [6–10].It is found that the support materials can strongly in fluence the activity of nickel catalysts.Several groups have reported that the nickel catalyst using TiO 2as the support is effective for CO methanation reaction [8,11,12],and the aim of the above studies is to remove carbon monoxide in hydrogen-rich gas for the use in polymer electrolyte fuel cells or ammonia synthesis.There are also some studies on methanation reaction using higher CO concentrations [13,14].However,it is well known that the reaction of CO methanation is a strongly exothermic reaction (3H 2+CO →CH 4+H 2O,ΔH°=−206kJ·mol −1)[6].When the nickel catalysts using conventional supports are used in the methanation reaction with a stream flow of H 2/CO=3,the reaction heat can rapidly accumulate onthe catalysts and make the catalysts sintered.Eisenlohr et al.reported that commercial methanation catalysts always showed a fast deactivation due to the temperature raise of catalyst bed [15].One way to solve the problem is to connect methanation reactors in series with intermediate gas cooling [1].In addition,the problem can also be solved by employing highly thermo-conductive and thermo-stable materials as the support of CO methanation catalysts.Silicon carbide (SiC)exhibits many superior properties,i.e.excellent thermostability,high mechanical strength,high chemical inertness,low coef ficient of thermal expansion and high thermal conductivity.Due to these properties,SiC could be employed as catalyst support in rigorous conditions,i.e.high endothermic or exothermic reactions,strong acidic or basic solution [16].Nguyen et al.[17]reported that SiC with medium surface area could function as catalyst support in the CO 2reforming of methane.Our group investigated the performance of Ni/SiC catalysts for the partial oxidation of methane and methane combustion,and found that the Ni/SiC catalysts showed high catalytic activity and stability [18,19].However,to our knowledge,nickel catalysts using SiC as the support are less investigated for CO methanation reaction.In this work,the catalytic behavior of Ni/SiC catalysts for CO methanation reaction with a stream flow of H 2/CO=3was investigated.The results showed that the silicon carbide supported nickel catalyst exhibited high catalytic activity and stability in CO methanation.2.Experimental 2.1.Catalyst preparationSiC support was prepared by a sol –gel and carbothermal reduction route [20],and TiO 2support was commercial product,P25.TheFuel Processing Technology 92(2011)2293–2298⁎Corresponding author.Tel.:+863514065282;fax:+863514050320.E-mail address:xyguo@ (X.-Y.Guo).0378-3820/$–see front matter ©2011Elsevier B.V.All rights reserved.doi:10.1016/j.fuproc.2011.08.002Contents lists available at ScienceDirectFuel Processing Technologyj o u r n a l h o me p a g e :w w w.e l s ev i e r.c om /l o c a t e /f u p ro ccatalysts were prepared by conventional impregnation method[18]. To prepare15wt.%Ni/SiC catalyst,1g of above SiC was added into 25.5mL Ni(NO3)2aqueous solution(0.1mol/L)with stirring for12h. Then the slurry was heated at80°C until nearly all the water evaporated and the mixture was dried at100°C for6h.Afterward the dried sample was calcined in air at different temperatures for4h. The similar processes can be used to prepare Ni/SiC catalysts with different nickel loadings.15wt.%Ni/TiO2catalyst was prepared by the similar route to15wt.%Ni/SiC catalyst.1g of TiO2was added into 25.5mL Ni(NO3)2aqueous solution and stirred for12h.After being heated and dried,the catalyst was calcined at550°C for4h in air.2.2.Catalyst testThe performance of the catalysts was tested in afixed-bed reactor (a40cm long stainless steel tube with an inner diameter of6mm). The reactor was heated up by a PID regulated oven and the reaction temperature was measured in the middle of the catalyst bed using a K-type thermocouple.Reaction gases,which consisted of H2and CO (molar ratio of H2/CO=3,without diluent gas),were supplied from high-pressure gas cylinders and theflow rate was controlled by mass-flow controller(MFC)to ensure a space velocity(GSV)of4500h−1.0.8mL of catalyst(sieve fraction40–60mesh)was placed in thecenter of the tubular reactor.All experiments were performed at a pressure of2.0MPa.Before each catalyst test,the catalyst was reduced at500°C in the CO/H2mixture gases for2h,then decreased to reaction temperature and kept for1h.The outlet gases were analyzed by GC-14B gas chromatograph with TDX-01column and a GDX-104 column using thermal conductivity detector andflame ionization detector.The selectivity of a certain product is calculated by the formula,S i=n i C i/∑n i C i,where n i and C i are the number of carbon atoms and the concentration of product“i”,respectively.2.3.Catalyst characterizationThe crystalline phases of the catalysts were analyzed by a Rigaku D-Max/RB X-ray diffractometer(XRD)with Cu Kαradiation with a scanning rate of6°/min.The catalyst morphology and structure were analyzed by a JEOL-2010transmission electron microscopy(TEM).The surface areas of catalysts were calculated from the BET method,which was performed at nitrogen temperature at77K on a Micromeritics Tristar3000analyzer.The nickel amount of catalysts was measured by ICP spectrometer.TG–DSC studies of the catalysts after reaction were performed on NETZSCHSTA409PC thermoanalyzer within a temper-ature range from room temperature to850°C at a heating rate of 10°C/min in airflow.3.Results and discussion3.1.CO methanation over Ni/SiC catalyst3.1.1.Influence of nickel loadingThe effect of metal loading on the activity of Ni/SiC catalysts was investigated,and the results are shown in Fig.1and Table1.It can be seen from Fig.1that the methanation activity of SiC supported catalysts increases with the nickel loading from5wt.%to15wt.%. Reaction rate of CO conversion which is defined as moles of CO converted per gram of Ni per second at250°C is shown in Table1.It can be seen that increasing the nickel loading from10wt.%to15wt.% results in an increase of the rate of CO conversion from1.28to 5.10μmol s−1g−1.However,when the nickel loading further in-creases to20wt.%,the rate of CO conversion at250°C decreases to 1.34μmol s−1g−1.As compared with15wt.%Ni/SiC catalyst,there is no significant increase in methanation activity for20wt.%Ni/SiC catalyst(Fig.1).The metal crystallite sizes calculated by Scherrer's equation[21]are shown in Table2.From the table,the metal crystallite size increases with increasing the nickel loading.The influences of the metal particle size on methanation activity have been studied by several authors.Panagiotopoulou et al.[8]reported that the rate of CO conversion increased by a factor of166with increasing the loading of Ru on TiO2from0.5wt.%to5wt.%.Takenaka et al.[11]reported that the Ni crystallites with relatively large sizes were more effective for CO methanation.Zhou et al.[22]demon-strated that the activity of CO hydrogenation increased with increasing the rhodium particle size in the range of 3.0–5.0nm. Therefore,CO methanation is a structure sensitive reaction and the larger particle size facilitates CO hydrogenation.20wt.%Ni/SiC catalyst has larger metallic particles but lower methanation activity than15wt.%Ni/SiC catalyst.The reason may be that both particle size and amount of active sites affect the methanation activity.It is well known that the larger crystallite size results in the decrease of metal dispersion on the surface of catalyst[8]and the latter one can affect the amount of active sites.Therefore,the lower rate of CO conversion may be due to the less active sites over20wt.%Ni/SiC compared with 15wt.%Ni/SiC catalyst.This is in accordance with the results reported by Aksoylu et al.[23],who found that the methane production per square meter of nickel surface area was enhanced with lower Ni loading over Ni/Al2O3catalysts.On the other hand,the large particle size on20wt.%Ni/SiC catalyst may block the external surface of support and this can also decrease the activity of CO methanation. Therefore,15wt.%is an optimal loading of Ni/SiC catalyst for methanation reaction.It should be noted that the products of CO hydrogenation under present reaction condition include methane,higher hydrocarbons (C2H6,C2H4,C3H8and C3H6),carbon dioxide and water.The forma-tion of CO2may be due to the water-gas-shift reaction,CO+Temperature (o C)EquilibriumCOConversion(%)T emperature (o C)COconversion(%)Fig.1.Influence of nickel loading on the activity of Ni/SiC catalysts and the calculated equilibrium CO conversions under P=2.0MPa,H2/CO=3and GSV=4500h−1.Table1Catalytic performance of Ni/SiC and Ni/TiO2catalysts.Catalyst Ni loading(wt.%)Rate of CO conversionat250°C(μmol s−1g−1)aCH4selectivityat320°C(%)Ni/SiC5–70.8910 1.2890.3115 5.1092.0420 1.3493.60Ni/TiO215 3.5282.21Reaction condition:H2/CO=3,GSV=4500h−1,and P=2.0MPa.a Converted CO per second per gram catalyst.2294Y.Yu et al./Fuel Processing Technology92(2011)2293–2298H2O=CO2+H2.The methane selectivity over the Ni/SiC catalysts at 320°C is shown in Table1.It can be seen that the methane selectivity increases with increasing the Ni loading.For CO methanation reaction, CO is adsorbed and dissociated on the surface of metal particles[24]and the dissociation of C\O bond in adsorbed CO species is the rate-determining step[25].It has been reported that there are three forms of adsorbed CO:linear CO,bridged CO and twin CO[26].The activity of adsorbed CO breaking into surface carbon and surface oxygen on the catalyst surface follows the sequence,bridged CO N linear CO N twin CO[10].According to the methanation mechanism,adsorbed CO is dissociated and converted into CH x as intermediate species by assistance of H on the surface of catalysts,and the concentration of the intermediate species determines the distribution of reaction products [25].For the methanation reaction,there are more active bridged CO on larger metallic particles[27],and then the rate of the dissociation and hydrogenation of intermediate species becomes faster[22,28]. Therefore,the methane selectivity is higher on high loading Ni/SiC catalysts.3.1.2.Thermodynamics and equilibrium conversionFrom Fig.1,the conversion of CO over Ni/SiC catalysts can increase up to100%with increasing the reaction temperature.When the reaction temperature is higher than440°C,however the CO con-version begins to decrease.It is most likely due to thermodynamic limitation.According to the thermodynamic equilibrium,theΔG can become positive when the temperature is higher than530°C[29]. Therefore,high temperature is favorable to the stream reforming of methane—the reverse of methanation.Supposing that the only product of CO hydrocarbon is methane, thermodynamic equilibrium data at the temperature range from100 to500°C are calculated.The calculation is performed under the same operating conditions as our experiments,i.e.H2/CO=3,P=2MPa. From Fig.1,the complete conversion of CO can be obtained below 250°C,and then the conversion slightly decreases with increasing the reaction temperature.The CO conversion decreases to87%when the reaction temperature rises to500°C.These are in agreement with the experimental results.3.1.3.Effect of calcination temperatureThe sizes of metallic particles in15wt.%Ni/SiC catalysts under different calcination temperatures are shown in Table2.It can be seen that the particle size increases with increasing the calcination temperature.It has been established that the methanation reaction is structure sensitive and larger particle size facilitates the cleavage of C\O bond[8,22,27,30].Therefore,the catalysts calcined at higher temperatures have higher catalytic activity for CO methanation.Fig.2 shows the influences of the calcination temperature on the activity of 15wt.%Ni/SiC catalyst for CO methanation.It can be seen that the CO conversion increases with increasing the calcination temperature from450°C to550°C.However,there is a little decrease in the CO methanation activity over the Ni/SiC catalyst calcined at600°C.This is likely due to that oversize particles formed at the high temperature could result in a decrease of active sites.Therefore,550°C is an optimal calcination temperature for15wt.%Ni/SiC catalyst.3.2.Performance of Ni/SiC and Ni/TiO2catalystsAccording to literature,TiO2supported nickel catalysts are effective for the CO methanation[8,11,12].Therefore,15wt.%Ni/SiC and15wt.%Ni/TiO2catalysts were tested under the same conditions for comparison.It can be seen from Table1that15wt.%Ni/SiC catalyst is more active than15wt.%Ni/TiO2catalyst.Fig.3shows the evolution of CO conversion over Ni/SiC and Ni/TiO2catalysts at340°C.For the Ni/SiC catalyst,there is an activity rise in the initial stage of the methanation reaction and CO is nearly converted completely.When the reaction time is longer than40h the activity of Ni/SiC catalyst exhibits a slow drop and the CO conversion decreases from initial97% to91%after100h.The decrease of activity is due to the losing of nickel by the formation of nickel carbonyl.According to ICP analysis,the nickel amount of the Ni/SiC catalyst has decreased by16%after reaction for100h.For Ni/TiO2catalyst,the activity decreases rapidly and the CO conversion decreases from initial98%to below50%afterTable2Particle sizes of different Ni/SiC and Ni/TiO2catalysts.Catalyst Ni loading(wt.%)Calculationtemperature(°C)Metal crystallitesize(nm)aNi/SiC55508.61055015.31545017.450018.155019.060020.92055023.5 Ni/TiO215550–Used Ni/SiC1555021.6b Used Ni/TiO21555023.5ba Calculated from NiO(021)plane by Scherrer's equation.b Calculated from Ni(111)plane by Scherrer's equation.COconversion(%)Temperature(o C)Fig.2.Influence of calcination temperature on the activity of Ni/SiC catalysts under P=2.0MPa,H2/CO=3and GSV=4500h−1.COconversion(%)Time (h)parison of the catalytic performance between15wt.%Ni/SiC and15wt.% Ni/TiO2catalysts under T=340°C,P=2.0MPa,H2/CO=3and GSV=4500h−1.2295Y.Yu et al./Fuel Processing Technology92(2011)2293–2298100h reaction.These results indicate that the Ni/SiC catalyst exhibits an obviously better stability than the Ni/TiO 2catalyst.3.3.TG –DSC analysis of used Ni/SiC and Ni/TiO 2catalystsTG –DSC analysis was performed for the used Ni/SiC and Ni/TiO 2catalysts and the results are shown in Fig.4.From the TG pro file shown in Fig.4A,no weight loss but a viewable increase of weight can be observed,indicating that there are no carbon depositions over both catalysts.The weight increase over both catalysts is likely due to the oxidation of metallic Ni to NiO.As seen in Fig.4B,no remarkable exothermic peaks can be observed for the used Ni/SiC and Ni/TiO 2catalysts,indicating that the oxidation of deposited carbon on the used catalysts does not occur.This is in agreement with the results reported by Zhu et al.[31].Therefore,the decrease in the activity of the Ni/SiC catalyst for methanation reaction is not due to the carbon deposition.3.4.XRD resultsFig.5shows the XRD results of 15wt.%Ni/SiC and 15wt.%Ni/TiO 2catalysts before and after reaction at 340°C for 100h.In the XRD patterns of fresh Ni/SiC catalyst shown in Fig.5A,the diffraction peak at 2θ=43.3°is attributed to NiO (012)plane [18].According to Scherrer's equation [21],the average NiO crystallite size calculated from the peak is about 19.0nm.In the XRD patterns of fresh Ni/TiO 2catalyst,diffraction peaks attributed to NiTiO 3phase can be detected (Fig.5A).Rao et al.[32]reported that NiTiO 3was formed whenNi/TiO 2catalyst was calcined above 500°C.Therefore the NiTiO 3phase in the present XRD patterns may be formed during the calcination.In our experiments,the color of Ni/TiO 2catalyst can become light yellow,which is the color of NiTiO 3[33].From both XRD patterns shown in Fig.5B,no diffraction peak of carbon can be found,indicating that the carbon deposition on the two methanation catalysts can be ignored.This is in accordance with the results of TG –DSC analysis.The diffraction peak at 2θ=44.56°is attributed to metallic Ni (111)[18].Metallic nickel exists in both catalysts,indicating that the active phase for CO methanation is metallic nickel.For the used Ni/SiC catalyst,the mean nickel particle size calculated from Ni (111)plane by Scherrer's equation [21]is about 21.6nm.The nickel particle size only has a slight increment during the reaction,suggesting that no serious sintering has occurred on the Ni/SiC pared with the fresh Ni/TiO 2catalyst,the crystallite size of used Ni/TiO 2increased signi ficantly (particle size from 24.2to 30.3nm and from 26.6to 35.9nm for rutile and anatase,respectively).The increase of particle size may be due to the production of hot spots during the CO methanation,which possibly make TiO 2crystallites sintered.In addition,the speci fic surface areas of Ni/TiO 2catalyst decreased from 32.1to 16.7m 2/g after reaction at 340°C for 100h.Ruckenstein et al.[34]reported that TiO x could1002003004005006007008009092949698100102104Ab aS a m p l e w e i g h t (%)temperature(o C)temperature(o C)a---15 wt.% Ni/SiC b---15 wt.% Ni/TiO 2BD S C /(m W /m g )Fig.4.TG –DSC pro files of used 15wt%Ni/SiC and 15wt.%Ni/TiO 2catalysts.AI n t e n s i t y (c p s )2 Theta (o )2 Theta (o )BI n t e n s i t y (c p s )Fig.5.XRD patterns of 15wt.%Ni/SiC and 15wt.%Ni/TiO 2catalysts before (A)and after (B)methanation reaction at 340°C for 100h.(●)NiTiO 3;(■)anatase;(▲)rutile;(★)SiC;(♦)NiO;(○)Ni.2296Y.Yu et al./Fuel Processing Technology 92(2011)2293–2298migrate onto the surface of metal nickel particles due to the strong interaction of support –metal.During the methanation reaction,the TiO 2may amalgamate with the reduced Ni due to the strong metal –support interaction.This is not favorable for CO absorption and decomposition on metal Ni.Therefore,the Ni/TiO 2catalyst shows a declining activity and stability in the methanation reaction.3.5.TEM characterization of Ni/SiC catalystsTEM images of fresh and used 15wt.%Ni/SiC catalyst are shown in Fig.6.From the two TEM images,it can be seen that the metal particles are well distributed on the support surface.From the image of fresh Ni/SiC catalyst (Fig.6A),the size of NiO particles ranges from 10to 25nm.In Fig.6B,the size of nickel particles in the used catalyst is about 10–30nm.These results are in accordance with those derived from XRD analysis.From both XRD and TEM results,15wt.%Ni/SiC catalyst shows better stability in the methanation reaction due to the excellent thermal conductivity and thermostability of the SiC support.4.ConclusionThe synthesis of methane from syngas was investigated over Ni/SiC catalysts which were prepared by impregnation method.From the present work,15wt.%Ni/SiC catalyst calcined at 550°C exhibits excel-lent catalytic activity for CO pared with Ni/TiO 2catalyst,the Ni/SiC catalyst exhibits higher activity and better stability in the CO methanation due to the excellent thermostability and 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生命科学研究2009年慢病毒载体是目前应用最广泛的基因运载工具之一，在基因治疗研究和转基因动物的制备中已显示出其广阔的应用前景．慢病毒载体是一种反转录病毒载体，其中以人类免疫缺陷性病毒（HIV -1）载体研究最为深入．与传统的小鼠白血病病毒载体（MuLV ）偏爱整合入基因5′端相比，慢病毒载体均匀分布于基因组内，从而降低了其激活原癌基因的几率［1~3］．重组慢病毒载体既能感染分裂期细胞，也能感染非分裂期细胞［4］．它可携带较大的外源基因（约8kb 左右）并稳定整合和表达［5］，加之其诱发的宿主免疫反应相对较小［6］，使得重组慢病毒载体具有较为广阔的应用应用荧光实时定量PCR 方法检测重组慢病毒滴度及其感染效率马海燕，方彧聃，张敬之*（上海交通大学医学院，上海市儿童医院上海市医学遗传研究所，中国上海200040）摘要：慢病毒载体已经广泛应用于动物模型中基因治疗的研究和转基因动物的制备，而准确地测定重组慢病毒的滴度和感染效率是其关键步骤．通过荧光实时定量PCR 的方法定量分析重组慢病毒的颗粒数以及病毒的活性滴度，并以GFP 报告基因的方法作为对照来验证定量PCR 方法的准确性．研究结果显示，应用荧光实时定量PCR 法与GFP 报告基因法测定得到的病毒活性滴度成正相关，而且前者可以更加准确地测定病毒滴度和病毒感染效率．关键词：慢病毒载体；荧光实时定量PCR ；病毒滴度；整合拷贝数；感染效率中图分类号：Q331文献标识码：A文章编号：1007-7847（2009）05-0394-05A Novel Method for the Determination of Recombinant LentiviralTiter and Infectivity by qRT -PCRMA Hai -yan ，FANG Yu -dan ，ZHANG Jing -zhi *（School of Medicine ，Shanghai Jiaotong University ，Shanghai Children ’s Hospital ，Shanghai Institute of Medical Genetics ，Shanghai 200040，China ）收稿日期：2009-03-18；修回日期：2009-05-16基金项目：国家高技术研究发展计划项目（2007AA021206）；国家自然科学基金资助项目（30870943）；上海市自然科学基金资助项目（08ZR1412100）作者简介：马海燕（1983-），女，山东淄博人，硕士研究生，主要从事病毒载体在转基因动物制备中的应用研究；*通讯作者：张敬之（1959-），男，上海人，上海交通大学医学遗传研究所副教授，博士，主要从事分子病毒学研究，Tel ：021-********，E -mail ：********************.Abstract ：Lentiviral vector is being widely used in the study of gene therapy in animal models and in generating transgenic animals.However ，determination of lentiviral particles and their infectivity is essential before their being used.Such a requirement can be accurately achieved by qRT -PCR.Refered by infectious units got from GFP reporter assay ，it showed a positive correlation between the two approaches.A reliable ，accurate and rapid method is therefore established for the determination of the recombinant lentiviral titer and the infectivity.Key words ：lentiviral vector ；qRT -PCR ；viral titer ；integration copy number ；infectivity（Life Science Research ，2009，13（5）：394～398）第13卷第5期生命科学研究Vol．13No．52009年10月Life Science Research Oct.2009第5期前景．无论何种目的使用重组慢病毒，都有必要准确地检测病毒滴度．目前，重组慢病毒滴度的检测方法有：依赖于报告基因的GFP荧光检测法、检测慢病毒外壳蛋白p24的抗原-抗体法（ELISA 法）和检测其逆转录酶活性的酶学法等．这些方法多存在耗时、费力、检测成本较高、病毒用量多、不适于非报告基因载体等缺点．所以，建立一种快速、简便、准确的慢病毒滴度检测方法，是非常有必要的．在此，我们介绍一种利用实时定量PCR测定重组慢病毒滴度的方法．该方法通过在载体的长末端重复序列区（LTR）设计定量引物，利用荧光实时定量PCR测定重组慢病毒中LTR拷贝数来测定病毒颗粒数和有效感染的病毒颗粒数．通过与GFP报告基因测定法的比较验证，证明该测定方法准确可靠．并且，通过慢病毒颗粒数与实际有活力病毒滴度的比较，可以计算得到重组慢病毒感染效率．多次重复实验证明，该方法具有快速、准确的优点，非常适用于非报告基因载体的病毒滴度及其感染效率的测定．1材料与方法1.1材料组成慢病毒载体的3个质粒FUGW（即含eGFP基因的转基因质粒），△8.9（编码结构和非结构蛋白基因质粒）和VSVG（外壳蛋白质粒）由美国Marine Medical Center Research Institute王征宇博士惠赠；293T细胞（人胚肾细胞）购自美国ATCC（American Type Culture Collection）细胞库；ProFection购自Promega公司；胎牛血清（FBS）、培养液和Hanks液及Salmon Sperm DNA 购自Gibco BRL；STE配方为10mmol/L Tris pH 7.6，1mmol/L EDTA pH8.0，0.1mol/L NaCl．1.2FUGW病毒的制备当293T细胞长至70%饱和度时，用磷酸钙法转染，按照Promega公司的ProFection Kit说明书操作，其中FUGW15μg，△8.910μg，VSVG7.5μg，转染6h后换完全培养液（含10% FBS的DMEM），并在37℃，5%CO2培养约60h．上述病毒培养上清液经离心、过滤后，50000g 超速离心1.5h后弃上清，在病毒沉淀上加少量Hanks液，获得病毒浓缩液，-80℃保存待用．1.3病毒RNA的提取取2μL病毒浓缩液进行DNase处理，体系中加入5μL10×DNase I Buffer、2μL DNase I （5U/uL，Takara Bio Inc，Shiga，Japan）、2μL RNase inhabitor（Takara Bio Inc.，Shiga，Japan），用DEPC水定容至50μL反应体系，37℃45 min．DNase处理后的混合液，加入350μL STE、20μL10%SDS和5μL蛋白酶K（20g/L，AMRESCO，Solon，OH），56℃15min水解．最后等体积酚、氯仿抽提，两倍体积无水乙醇沉淀，冻干后，20μL DEPC水溶解．1.4反转录反应病毒RNA在酚/氯仿抽提、无水乙醇沉淀前，需经过DNase I处理，以避免DNA污染．根据产品说明书，取1μg RNA、20pmol RT-PCR 下游引物（5′-GAGAGCTCCCAGGCTCAGATC-3′）、2μL5×RT Buffer、1μL MLV酶（Takara Bio Inc，Shiga，Japan）、水补足至10μL体系，37℃1h．1.5实时定量PCR分析病毒颗粒数为了测定制备的慢病毒的病毒颗粒数，应用实时定量PCR测定病毒LTR拷贝数.其中，引物和探针序列见表1．在反应体系中，引物900 nmol/L，探针250nmol/L，2.5μL10×Ex-Buffer，15μmol Mg2+，2.5μmol dNTP，1U ExTaqE，5μL样本，总反应体系为25μL体积．实时定量PCR仪（Corbett Life Science RG-3000，Sidney，Australia）上反应：95℃5min变性，95℃30s，59℃30s，40个循环，59℃520nm处检测荧光值．软件分析荧光检测数据．1.6不同剂量FUGW病毒感染293T细胞取病毒浓缩液，按10倍稀释法，取0.1、0.01、0.001μL病毒浓缩液（即用实时定量PCR 检测法，定量慢病毒颗粒数为：6.32×107、6.32×106、6.32×105），用含有8mg/L Polybrene促感染（Sigma-Aldrich，Inc，St.Louis，MO）且不含血清的培养液逐级稀释后，感染293T细胞（1.5×106/孔），37℃2h.然后，加入完全培养液培养细胞2d，表1实时定量PCR引物及探针序列Table1The sequences of primer and probe of Real-time PCRLTR-FLTR-PLTR-ProbePrimer Sequences5′-ACAGCCGCCTAGCATTTCAT-3′5′-GAGAGCTCCCAGGCTCAGATC-3′5′-ACATGGCCCGAGAGCTGCATCC-3′马海燕等：应用荧光实时定量PCR方法检测重组慢病毒滴度及其感染效率395生命科学研究2009年图1定量PCR 反应荧光强度曲线Fig.1Fluorescence intensity curve of Real -time PCR图2定量PCR 标准曲线Fig.2Standard curve of Real -time PCR至荧光显微镜下观测绿色荧光蛋白表达情况．1.7病毒感染细胞内DNA 的提取细胞经2d 培养后，用胰酶将细胞消化，收集入1.5mL EP tube 中，200g ，5min ，将细胞沉淀下来．加入200μL STE 、20μL 10%SDS 、10μL 蛋白酶K ，混匀后，37℃4h.加入等体积酚，振荡混匀，15000g ，离心12min ；吸取上清，加入等体积的氯仿，振荡混匀，15000g 离心6min ；吸取上清，加入1/10体积3mol/mL NaAc ，两倍体积的无水乙醇，-20℃沉淀1h 以上；混合液15000g ，4℃离心20min ；弃上清，沉淀风干，100μL TE 溶解．1.8实时定量PCR 测定重组载体整合拷贝数为了测定被感染的293T 细胞内重组载体的整合，用实时定量PCR 检测上述被抽提的基因组DNA 中LTR 的拷贝数．具体方法同1.5．2结果2.1实时定量PCR 检测病毒颗粒数本实验中，定量PCR 引物设计在LTR 区，由于一个慢病毒含有两个病毒基因拷贝，因此，在计算病毒颗粒数时，LTR 拷贝数除以2即为病毒颗粒数.应用实时定量PCR 检测到病毒LTR 拷贝数为1.26×1012/mL ，定量PCR 反应荧光强度曲线及标准曲线见图1、图2.通过计算，得到病毒颗粒数为6.32×1011/mL ．此时计算得到的病毒颗粒数为所有收集到的病毒颗粒总数，既包括有感染效力的病毒，也包括无感染效力的病毒．为了测定制备得到的病毒实际滴度（即单位体积内有感染效力的病毒颗粒数），我们将不同剂量病毒感染细胞，分别用GFP 报告基因法和定量PCR 法测定病毒滴度．2.2实时定量PCR 检测病毒载体整合拷贝数，计算病毒滴度取病毒浓缩液，按照10倍稀释，分别取0.1、0.01、0.001μL 病毒浓缩液感染1.5×106293T 细胞，感染2d 后，应用实时定量PCR 检测不同病毒量感染的细胞DNA 中LTR 整合拷贝数，结果显示，0.1、0.01、0.001μL 病毒感染的细胞中外源基因整合的拷贝数分别为5.32×106、9.28×105、4.48×104.定量PCR 的系统参数为：R =0.99，R ∧2=0.99，Efficiency =1.01，各参数值表明实验检测的准确性和可信性．当一个病毒感染细胞并将外源基因整合入基因组后，由于其末端发生跳跃过程，使得每条DNA 单链上含有两个完整的LTR ，因此，在被整合的细胞基因组，一个慢病毒颗粒=LTR 拷贝数/4．通过计算，得到病毒滴度为（1.59±0.64）×1010IU /mL ．2.3GFP 报告基因检测法测定病毒滴度同时将0.1、0.01、0.001μL 病毒浓缩液感染2d 后的细胞，置于荧光显微镜下观测．镜检结果显示，GFP 阳性细胞数逐级递减，0.1、0.01、0.001μL 病毒量感染的细胞中，GFP 阳性率分别为52%、6%、0.5%，呈现较好的倍比关系（图3）．通过公式：病毒滴度=感染细胞数×GFP 阳性率×病毒稀释倍数÷病毒量，得出病毒滴度为（8.10±0.79）×109IU/mL ．2.4GFP 报告基因检测法与定量PCR 检测法所得病毒滴度比较，验证定量PCR 检测法准确性通过比较GFP 报告基因检测法和实时定量510152025303510^-110^-210^-3N o r m f l u o r eCycle numberThreshold·····10^410^510^610^710^810^940302010R =0.99980R ^2=0.99960Efficiency=0.96M =0.293B =13.087Concentration （Copy number ）T h r e s h o l d c y c l e396第5期注：病毒感染293T 细胞数为1.5×106.Notes ：293T cells in each well are:1.5×106.图3逐级稀释的慢病毒感染293T 细胞后GFP 的表达情况（A ）0.1μL 病毒感染细胞；（B ）0.01μL 病毒感染细胞；（C ）0.001μL 病毒感染细胞.Fig.3GFP expression in infected 293T cells after 10fold dilution （A ）293T cells infected by 0.1μL virus ；（B ）293T cells infected by 0.01μL virus ；（C ）293T cells infected by 0.001μL virus.PCR 检测法（表2），可以看出，与我们预期结果一致，实时定量PCR 法检测结果与GFP 报告基因检测法检测结果成正相关．多次重复实验结果均表明，两者的检测结果成稳定的正相关性．而且，实时定量PCR 检测法不依赖GFP 蛋白正常表达，对整合入宿主基因组但不能正常表达的病毒仍然能够在其检测范围之内，因此，实时定量PCR 的检测结果更接近实际值，能够更加准确的检测病毒滴度．（A ）（B ）（C）表2GFP 报告基因检测法与定量PCR 检测法测得病毒滴度比较Table 2Comparison of lentivirus titer determined by GFP reporter assay and qRT -PCR approachDose of viral particles6.3×107 6.3×106 6.3×105GFP reporter assay Percentage of GFP +cells Infectious units qRT -PCR assayInfectious units52.0%7.8×1051.3×1066.0%9.0×1042.3×1050.5%7.5×1031.1×1042.5荧光实时定量PCR 法测定病毒实际感染效率通过计算公式：感染效率=有效感染病毒数/感染细胞总病毒颗粒数×100%，得到该实验中慢病毒的感染效率为：2.65±1.07%．3讨论慢病毒载体的研究，目前以HIV -1最为深入．慢病毒载体构建的基本原理是将HIV -1基因组中的基本骨架与编码其功能蛋白相分离，分别改建成载体质粒和表达包装蛋白的质粒，并将两种成分共转染入细胞，从中获得只有一次感染能力而没有复制能力的HIV -1载体假病毒［7］，从而提高了其应用的安全性.近年来，越来越多的研究者利用慢病毒载体系统作为在动物模型研究基因治疗的导入系统，并取得良好的效果［8~10］．与此同时，利用慢病毒载体介导制备转基因动物的研究也得到发展，慢病毒载体介导成功制备了转基因小鼠［11，12］、转基因猪［13］、转基因牛［14］等动物，为基因工程领域的研究奠定基础．由于慢病毒载体的基因转导效率主要取决于病毒滴度，这就使得慢病毒滴度及其感染效率的检测变得很重要．目前，通行的慢病毒滴度检测方法有：1）p24等抗原酶联检测法（p24ELISA 方法）.其缺点是：商业化的ELISA 试剂盒往往太贵，约6000元人民币一盒.而且蛋白含量检测结果不能直接反映其拷贝数；2）使用报告基因系统，无法检测其真实颗粒数及不携带报告基因的假病毒滴度；3）检测逆转录酶活性，用量马海燕等：应用荧光实时定量PCR 方法检测重组慢病毒滴度及其感染效率397生命科学研究2009年大，操作复杂及准确性差．本文所阐述的通过实时定量PCR法测定LTR拷贝数来检测病毒滴度的方法，能在提高慢病毒滴度检测准确性的同时，缩短检测时间、减少检测成本.相比传统检测方法，实时定量PCR 检测法有以下特点：1）由于本检测方法中，实时定量PCR的检测引物设计在慢病毒载体的LTR区，因此检测不依赖于所携带的外源基因；2）传统的GFP报告基因检测方法依赖绿色荧光蛋白的表达，对于外源基因整合入宿主基因组中但由于基因沉默而未能表达GFP蛋白的细胞无法检测，致使滴度测定不能准确地反应病毒的感染效率；而实时定量PCR检测法不依赖GFP报告基因的功能表达，因此其准确程度更高；3）报告基因检测法，因其感染和表达效率随宿主细胞而异，而利用实时定量PCR方法，直接检测病毒的颗粒数和整合入宿主基因组内的外源基因的拷贝数，从而大大提高了其检测的准确性．目前，临床上及实验室所应用的定量PCR方法，通常是检测整合入宿主细胞的基因拷贝数，因此，是对有活力病毒滴度的测定．而本文所介绍的方法，是通过直接裂解病毒，利用荧光实时定量PCR检测总的病毒颗粒数，并结合传统的慢病毒活力滴度的检测方法，对病毒颗粒的感染效率进行检测．所以本方法更适于被应用于研究病毒制备、感染过程中各因素对病毒感染效率的影响．病毒浓缩和感染过程中，由于受超离的压力、反复冻融、受体细胞易感性、病毒自身半衰期等诸多因素影响，使得有效的病毒数要低于其总颗粒数．为了确定实验中病毒的用量，需要预测病毒的实际感染效率．利用本文所介绍的实时定量PCR方法，检测慢病毒颗粒数及有活性的病毒滴度，通过计算有活力的病毒颗粒和总病毒颗粒的比值，我们可以得到每次制备的病毒的感染效率．在我们的实验中，所得到的病毒在293T细胞的实际感染效率为4%左右．而且多次实验表明，反复冻融对病毒感染效率具有很大影响.病毒实际感染效率的测定，为我们在进行具体实验中确定病毒用量具有实际指导意义．经本实验室多次重复试验，结果均表明荧光实时定量PCR法是一种高效、准确、快速的检测重组慢病毒滴度及其感染效率的方法，为重组慢病毒载体的应用奠定了基础．致谢：感谢任兆瑞教授对本文的悉心指导．参考文献（References）:[1]WU X，LI Y，CRISE B，et al.Transcription start regions inthe human genome are favored targets for MLV integration[J].Science，2003，300：1749-1751.[2]DEPALMA M，MONTINI 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关于中国天眼的英文作文英文回答：The Five-hundred-meter Aperture Spherical Telescope (FAST), also known as China's "Eye of the Sky," is a colossal astronomical marvel that has revolutionized the study of the cosmos. Situated in a remote karst depression in Pingtang County, Guizhou Province, FAST is the world's largest and most sensitive radio telescope, surpassing the Arecibo Observatory in Puerto Rico.With a massive dish measuring an impressive 500 meters in diameter, FAST boasts an enormous collecting area that gives it unparalleled sensitivity and resolving power. Its design is based on a novel concept known as the "active surface," where the entire dish surface is adjustable, enabling it to focus on specific regions of the sky with high precision.FAST's construction, spanning over a decade, was atestament to China's technological prowess and scientific ambitions. The telescope began scientific operations in September 2016 and has since made numerous groundbreaking discoveries. Among its most notable contributions are the detection of pulsars, the study of interstellar gas, and the investigation of extragalactic objects.One of FAST's most significant achievements is its discovery of an enormous number of millisecond pulsars, rapidly rotating neutron stars that emit precise pulses of radio waves. These pulsars serve as invaluable probes for studying the evolution of binary systems, testing theories of gravity, and exploring the enigmatic nature of dark matter.FAST has also played a crucial role in mapping the distribution of interstellar gas, the raw material from which stars form. By observing the faint hydrogen radiation emitted by these gas clouds, astronomers have gained insights into the formation and evolution of galaxies, including our own Milky Way.Moreover, FAST has extended our astronomical reach beyond our galaxy, enabling the detection and characterization of extragalactic objects. Its sharp resolution has allowed astronomers to probe the structure and dynamics of distant galaxies, unveil the nature ofblack holes, and search for signs of extraterrestrial life.The scientific community worldwide has embraced FAST as a valuable asset, providing access to this cutting-edge instrument through open calls for observing proposals. Researchers from various countries have utilized FAST's capabilities to pursue transformative research projects, leading to advancements in our understanding of the universe.中文回答：中国天眼又称“天眼”，坐落在贵州平塘县的一个喀斯特洼地，是世界上最大、最灵敏的射电望远镜，超过了波多黎各的阿雷西博天文台。

ANDREW S. TANENBAUM 秒，约533 msec.----- COMPUTER NETWORKS FOURTH EDITION PROBLEM SOLUTIONS 8. A collection of five routers is to be conn ected in a poi nt-to-poi nt sub net.Collected and Modified By Yan Zhe nXing, Mail To: Betwee n each pair of routers, the desig ners may put a high-speed line, aClassify: E aEasy, M ^Middle, H Hard , DaDeleteGree n: Importa nt Red: Master Blue: VI Others:Know Grey:—Unnecessary ----------------------------------------------------------------------------------------------ML V Chapter 1 In troductio nProblems2. An alter native to a LAN is simply a big timeshari ng system with termi nals forall users. Give two adva ntages of a clie nt-server system using a LAN.(M)使用局域网模型可以容易地增加节点。

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人脸识别技术论文人脸识别，特指利用人脸视觉特征信息的分析比较结果进行身份鉴别的计算机技术。

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人脸识别技术论文篇一人脸识别技术综述摘要：文章首先对人脸识别技术进行了介绍，其次回顾了人脸识别研究的发展历程及识别方法的基本分类，然后对当前主流的人脸识别方法展开了详细的论述，最后提出了人脸识别技术面临的问题及研究方向。

关键词：人脸识别;特征脸;线形判别分析;局部二值模式中图分类号：TP391Survey of face recognition technologyHe Chun(Education and Information Technology Center， China West Normal University， Nanchong Sichuan 637002， China) Abstract：This paper introduces technology of face recognition firstly， and reviews the development process and the basic classification method of face recognition. After that，the paper discusses the current methods of face recognition in detail， therefore proposes the existing problems in the research of recognition faces and future’s research direction.Key words：face recognition; Eigenface; linear discrimination analysis; LBP1 人脸识别技术简介人脸识别，特指利用人脸视觉特征信息的分析比较结果进行身份鉴别的计算机技术[1]。