卡介苗中具启动子活性片段克隆及分析

利用启动子探针型质粒ｐＫＫ２３２－８从卡介苗染色体ＤＡＮ的ＢａｍＨＩ酶切片段中克隆到４个具启动功能的ＤＮＡ片段,测定各重组子对氯霉素（Ｃｍ）的抗性,其中含３．３Ｋｂ外源片段的重组质粒ｐＢＣＧ２大肠杆菌转化子在ＬＭ培养基上对Ｃｍ抗性可达１７０×１０－６ｇ／ｍＬ,作了该片段的限制性酶切图谱．对ｐＢＣＧ２利用ＳａｌⅠ位点,亚克隆出４种部分重叠的具启动功能的ＤＮＡ片段,这４种重组质粒分别命名为ｐＢＣＧ２－１,ｐＢＣＧ２－２,ｐＢＣＧ２－６和ｐＢＣＧ２－８,并分析了其中插入片段的大小及其转化子对Ｃｍ的抗性．将ｐＢＣＧ２－２的ＳａｌⅠ外源片段插入到分枝杆菌启动子探针型穿梭质粒ｐＥＱ３,获得一个可在卡介苗中表达ｌａｃＺ基因的重组子ｐＥＢ２２．斑点杂交实验证明,具有启动功能的ＤＮＡ片段来源于卡介苗的染色体ＤＮＡ．结果表明克隆到一个对大肠杆菌和卡介苗均具启动子活性的ＤＮＡ片段     

棒状杆菌表达载体pJL23的构建

质粒ｐＧＡ４６－４带有从谷氨酸棒杆菌染色体上分离到的有启动功能的片段,用ＰＣＲ技术从ｐＧＡ４６－４中扩增了该片段的关键区域;启动子ＰＧＬ,将该启动子经ＥｃｏＲⅠ－ＢａｍＨ Ⅰ双酶切后,与大肠杆菌质粒ｐＪＬ０１的ＥｃｏＲⅠ－ＢａｍＨⅠ大片段连接,再接入Ｘｙｌ Ｅ ｇｅｎｅ和棒状杆菌质粒ｐＸＺ１０１４２,构建成棒状杆菌－大肠杆菌穿梭表达载体ｐＪＬ２３。用邻苯二酚双加氧酶基因检测表明,该表达载体可在棒状     

抗砷质粒pSDR941的构建及其在专性自养极端嗜酸性…

将具有广泛寄主范围的ＩｎｃＱ质粒ｐＪＲＤ２１５的Ｋｍ基因片段删掉，置换进大肠杆菌抗砷质粒ｐＵＭ３上的抗砷基因及Ｐ１启动子片段，构建成新的抗砷质粒ｐＳＤＲ９４１，为１２．４ｋｂ，通过接合的方式将其转移到氧化硫硫杆菌中并获得了表达。     

大肠杆菌-分枝杆菌穿梭表达质粒PY-α的构建

利用分子生物学方法，以含人结核杆菌热休克蛋白（HSP）70基因全长序列的质粒pMT-70为模板，扩增出hsp70启动子，将其定向克隆于pUC-19，构建成重组质粒pY6013；然后以重组质粒pIJK-1为模板，扩增出分枝杆菌α信号肽基因并将其克隆到pY6013闰中hsp70启动子的下游，得到了大肠杆菌－分枝杆菌表达质粒pY-α。     

透明颤菌血红蛋白基因在苏云金杆菌中的表达

采用大肠杆菌与枯草杆菌的穿梭质粒pMK4作为载体,构建了含有果聚糖酶基因(sacB)启动子和vgb基因的重组质粒pMK-SV.通过原生质体和电转化,将pMK-SV转入苏云金杆菌中,经影印筛选得到转化子Bt4.对Bt4诱导表达,与原菌株相比Bt4的生长量增加了20.2%.     

抗砷质粒pSDR941的构建及其在专性自养极端嗜酸性氧化硫硫杆菌中的表达

将具有广泛寄主范围的ＩｎｃＱ质粒ｐＪＲＤ２１５的Ｋｍｒ基因片段删掉，置换进大肠杆菌抗砷质粒ｐＵＭ３上的抗砷基因及Ｐ１启动子片段，构建成新的抗砷质粒ｐＳＤＲ９４１，为１２．４ｋｂ，通过接合的方式将其转移到氧化硫硫杆菌中并获得了表达，与对照菌相比，含质粒ｐＳＤＲ９４１的氧化硫硫杆菌的抗砷水平从１５ｍｍｏｌ／Ｌ提高到３０ｍｍｏｌ／Ｌ．     

PheAB基因在棒状杆菌染色体上的整合

改造大肠杆菌质粒ｐＬＣＸ３１,切除其中的ｘｙｌＥ基因得到大肠杆菌质粒ｐＪＬ０１．将源于大肠杆菌分枝酸变位酶－预苯酸脱水酶基因２．３ｋｂ ＢａｍＨＩ片段克隆到大肠杆菌质粒ｐＪＬ０１中启动子Ｐ３２的下降,构建成质粒ｐＪＬ０２。再在ｐＪＬ０２的ＨｉｎｄⅢ位点接入棒状杆菌染色体的ＨｉｎｄⅢ消化的随机片段,构建成带不同棒状杆菌染色体片段而不带棒状杆菌自主复制顺序的棒状杆菌整合质粒ｐＪＬ０３。     

三种新的穿梭质粒pCC10、pCCT7和pST16的构建和性质研究

利用大肠杆菌质粒pUC18、pTG206和金黄色葡萄球菌质粒pC194在体外构建3个嵌合质粒pCC10、pCCT7和pST16.3个新质粒均是大肠杆菌和枯草杆菌的穿梭质粒。它们在大肠杆菌中呈Ap~rCm~r型,在枯草杆菌中则为Cm~rAp~5型。其中pCCT7和pST16含有xylE基因,可作为启动子探测质粒。所有的新质粒都具有来自pUC18的多酶克隆位点,适合于在大肠杆菌和枯草杆菌中重组外源DNA以及比较它们的表达情况。     

运用绿色荧光蛋白探讨肉葡萄球菌双精氨酸分泌途径

根据肉葡萄球菌(Staphylococcus carnosus)TM300的基因组序列设计引物,经PCR扩增得到其tufA基因的启动子Peftu片段与大肠杆菌–葡萄球菌穿梭载体pBT2-Tat-GFP连接,构建了穿梭质粒pBT2-ETG.结果表明,穿梭质粒pBT2-ETG成功地转入大肠杆菌与肉葡萄球菌宿主中稳定表达具有活性的绿色荧光蛋白GFP,并被转运到肉葡萄球菌宿主的细胞壁,为进一步研究肉葡萄球菌双精氨酸(Tat)转运系统正确分泌其他外源蛋白奠定了实验基础.     

r结核分枝杆菌PhoP／PhoR基因重组穿梭表达质粒的构建及鉴定

为构建结核杆菌PhoPR双组分系统PhoP、PhoR基因的重组穿梭质粒pMV361-PhoP、pMV361-PhoR并对其进行鉴定。采用结核分枝杆菌国际标准强毒株H37Rv菌株基因组DNA为模版,利用PCR技术分别扩增PhoP、PhoR基因编码序列,先克隆于T—Vector pMD19（Simple）,再亚克隆至大肠杆菌-分枝杆菌穿梭表达载体pMV361,并转化E.coli DH5α感受态细胞,提取所得的阳性克隆子获得重组表达质粒pMV361-PhoP、pMV361-PhoR,通过PCR及双酶切鉴定。结果显示,结核分枝杆菌国际标准强毒株H37Rv菌株基因组DNA中扩增出的PhoP、PhoR基因与GenBank公布的序列一致,经过PCR及双酶切鉴定,表明PhoP、PhoR基因均成功插入了pMV361穿梭载体。由此可知,成功构建了重组穿梭质粒pMV361-PhoP、pMV361-PhoR,为进一步研究PhoPR双组分系统奠定了基础。     

Markedness and its Interpretation in Language Use

Language markedness is a common phenomenon in languages, and is reflected from hearing, vision and sense, i.e. the variation in the three aspects such as phonology, morphology and semantics. This paper focuses on the interpretation of markedness in language use following the three perspectives, i.e. pragmatic interpretation, psychological interpretation and cognitive interpretation, with an aim to define the function of markedness.     

Features of Women's language

Language is a means of verbal communication. People use language to communicate with each other. In the society, no two speakers are exactly alike in the way of speaking. Some differences are due to age, gender, statue and personality. Above all, gender is one of the obvious reasons. The writer of this paper tries to describe the features of women＇s language from these perspectives： pronunciation, intonation, diction, subjects, grammar and discourse. From the discussion of the features of women＇s language, more attention should be paid to language use in social context. What＇s more, the linguistic phenomena in a speaking community can be understood more thoroughly.     

An Analysis of Catherine＇s Character——Wuthering Heights

Wuthering Heights, Emily Bronte＇s only novel, was published in December of 1847 under the pseudonym Ellis Bell. The book did not gain immediate success, but it is now thought one of the finest novels in the English language. Catherine is the key character of this masterpiece, because everybody and everything center on her though she had a short life. We can understand this masterpiece better if we know Catherine well.     

Lower Paleozoic oil relationships within Williston Basin, Canada

The Williston Basin is a significant petroleum province, containing oil production zones that include the Middle Cambrian to Lower Ordovician, Upper Ordovician, Middle Devonian, Upper Devonian and Mississippian and within the Jurassic and Cretaceous. The oils of the Williston Basin exhibit a wide range of geochemical characteristics defined as "oil families", although the geochemical signature of the Cambrian Deadwood Formation and Lower Ordovician Winnipeg reservoired oils does not match any "oil family". Despite their close stratigraphic proximity, it is evident that the oils of the Lower Palaeozoic within the Williston Basin are distinct. This suggests the presence of a new "oil family" within the Williston Basin. Diagnostic geochemical signatures occur in the gasoline range chromatograms, within saturate fraction gas chromatograms and biomarker fingerprints. However, some of the established criteria and cross-plots that are currently used to segregate oils into distinct genetic families within the basin do not always meet with success, particularly when applied to the Lower Palaeozoic oils of the Deadwood and Winnipeg Formation.     

低渗透非均质砂岩油藏启动压力梯度研究

理论推导与室内实验相结合,建立了低渗透非均质砂岩油藏启动压力梯度确定方法。首先借助油藏流场与电场相似的原理,推导了非均质砂岩油藏启动压力梯度计算公式。其次基于稳定流实验方法,建立了非均质砂岩油藏启动压力梯度测试方法。结果表明:低渗透非均质砂岩油藏的启动压力梯度确定遵循两个等效原则。平面非均质油藏的启动压力梯度等于各级渗透率段的启动压力梯度关于长度的加权平均;纵向非均质油藏的启动压力梯度等于各渗透率层的启动压力梯度关于渗透率与渗流面积乘积的加权平均。研究成果可用于有效指导低渗透非均质砂岩油藏的合理井距确定,促进该类油藏的高效开发。     

On Tragic Consciousness In The Works By Ernest Hemingway

As an American modern novelist who were famous in the literary world, Hemingway was not a person who always followed the trend but a sharp observer. At the same time, he was a tragedy maestro, he paid great attention on existence, fate and end-result. The dramatis personae's tragedy of his works was an extreme limit by all means tragedy on the meaning of fearless challenge that failed. The beauty of tragedy was not produced on the destruction of life, but now this kind of value was in the impact activity. They performed for the reader about the tragedy on challenging for the limit and the death.     

Design and analysis of generic LBS application developing platform

Location based services is promising due to its novel working style and contents.A software platform is proposed to provide application programs of typical location based services and support new applications developing efficiently. The analysis shows that this scheme is easy implemented, low cost and adapt to all kinds of mobile nework system.     

Achieving Unusual States of Matter Under High Pressure

正The periodicity of the elements and the non-reactivity of the inner-shell electrons are two related principles of chemistry,rooted in the atomic shell structure.Within compounds,Group I elements,for example,invariably assume the+1 oxidation state,and their chemical properties differ completely from those of the p-block elements.These general rules govern our understanding of chemical structures and reactions.Using first principles calcula-     

Exchange-Correlation and Electronic Excitation Energies from Pairing Matrix Fluctuations

We have developed an adiabatic connection to formulate the ground-state exchange-correlation energy in terms of pairing matrix linear fluctuations.This formulation of the exchange-correlation energy opens a new channel for density functional approximations based on the many-body perturbation theory.We illustrate the potential of such approaches with an approximation based on the particle-particle Random Phase Approximation(pp-RPA).This re-     

Density Functional Theory for the Response of Periodic Systems to Electric Fields Based on the Vector Potential Approach

正The electronic and nuclear(structural/vibrational)response of 1D-3D nanoscale systems to electric fields gives rise to a host of optical,mechanical,spectral,etc.properties that are of high theoretical and applied interest.Due to the computational difficulty of treating such large systems it is convenient to model them as infinite and periodic(at least,in first approximation).The fundamental theoretical/computational problem in doing so is that