重组人Endostatin在甲醇酵母Pichia Pastoris中的分泌表达

为了获得高效表达分泌型重组人Endostatin菌株 ,利用基因工程技术 ,将目的基因连接到Pichia表达载体 pPIC9K中 ,将该重组质粒转入GS115菌株后筛选His阳性菌株 ,并用G4 18筛选高拷贝阳性克隆菌 .重组表达的Endostatin经SDS PAGE电泳 ,可在分子量 2 2kD处见到单一目标带 .经体外生物学活性检测 ,Pichia菌株表达的Endostatin能有效地抑制培养的ECV30 4细胞及P6 1细胞的生长 .     

规模化鸡场粪样和苍蝇产CMY-2大肠杆菌中耐药基因与毒力基因调查及其共转移研究

为了解全国20家规模化鸡场粪样和苍蝇产CMY-2大肠杆菌中耐药基因与毒力基因的流行现状及其共转移情况,对2013-2015年间分离自粪样和苍蝇中的549株大肠杆菌,采用聚合酶链反应（PCR）方法筛选出blaCMY-2阳性菌株;脉冲场凝胶电泳（PFGE）分析blaCMY-2阳性菌株的遗传进化关系;K-B纸片法检测阳性菌株对13种抗菌药物的耐药性;利用PCR技术检测19种相关耐药基因以及14种毒力基因;接合转移试验和质粒复制子分型研究耐药基因及毒力基因的传播方式。结果,33株大肠杆菌呈CMY-2阳性,阳性率为6.0 %,且均表现为多重耐药;检测到15种耐药基因（blaTEM-1、blaCTX-M-55、blaOXA-1、qnrBDS、aac(6’)-Ib-cr、oqxA、tetA、tetC、sul1、sul2、sul3、rmtB和flor）和4种毒力基因（iutA、traT、fyuA和VagC）;33株携带blaCMY-2基因菌株中有21株接合转移成功,blaCMY-2基因可通过lncA/C或lncI1质粒与耐药基因（blaTEM-1、blaCTX-M-55、tetA或sul2）和（或）毒力基因（traT或VagC）共同转移。本研究阐明了规模化鸡场产CMY-2大肠杆菌携带耐药基因与毒力基因的情况,证实了细菌中质粒介导的耐药与毒力的共转移,为耐药性传播及疾病防控提供了参考。     

小肠结肠炎耶氏菌体外毒力试验

为了初步研究小肠结肠炎耶氏菌（Ｙ．ｅ．）致病机理，本文报告应用依钙、自凝、刚果红平板毒力试验和质粒ＤＮＡ提取方法，进行Ｙ．ｅ．体外毒力探讨。结果表明：带毒力质粒的Ｙ．ｅ．菌株Ｇ１５１，９２０和ＬＡＢ－Ｂ１８２依钙、自凝和刚果红平板毒力试验阳性；不带毒力质粒的Ｙ．ｅ．菌株８２－１４０和４３９－８０Ｖ￣－依钙、自凝和刚果红平板毒力试验阴性。依钙、自凝、刚果红平板毒力试验阳性与携带毒力质粒的一致性以及依钙、自凝、刚果红平板毒力试验阴性与不携带毒力质粒的一致性表明：Ｙ．ｅ．的依钙、自凝和吸收刚果红特性是毒力质粒编码。     

深黄被孢霉△6-脂肪酸脱氢酶基因敲除及鉴定

以产油丝状真菌深黄被孢霉M6-22(Mortierella isabellina 6-22)为出发菌株,利用亚硝基胍诱变技术筛选到一株对潮霉素药物敏感的菌株Mortierella isabellina 6-22-4,同时构建了含有潮霉素抗性基因表达单元和深黄被孢霉△6-脂肪酸脱氢酶基因5'与3'端部分同源序列的重组质粒PD4MI6,用于对深黄被孢霉△6-脂肪酸脱氢酶基鼠的敲除.通过PEG/CaCl2介导的原生质体转化法,将质粒PD4MI6转化入深黄被孢霉M6-22-4菌株.转化子经潮毒素抗性选择平板筛选、分子检测获得了一个△6-脂肪酸脱氢酶基因缺失的突变株Mut6.气相色谱分析结果显示,该突变株γ-亚麻酸特征峰消失,相应地亚油酸产量显著增加.培养结果表明该突变株的菌落形态、颜色及生长状况与出发菌株M6-22或M6-22-4相比均有显著改变.     

柞栎象布氏白僵菌优良菌株筛选

探索环境友好的柞栎象长效控制技术,筛选防治柞栎象效果较好的优良菌株,为柞栎象的生物防治和高效制剂研制奠定基础.对16株来源于自然侵染柞栎象幼虫、其他昆虫和利用柞栎象幼虫人工诱集的白僵菌菌株进行形态鉴定和生物学、毒力测定;综合评价菌株生长速率、产孢量、孢子萌发率和毒力4个因子的特征值,采用独立淘汰法和综合秩次选择法联合筛选.结果表明:16株白僵菌均为布氏白僵菌,筛选出Bbbh1、Bbbh9、Bbbh13和Bbbh16共4株优良菌株.     

丢失毒力质粒的小肠结肠炎耶氏菌实验性感染家兔致病机理研究

本文报告应用致病性血清型丢失毒力质粒Ｙ．ｅ菌株４３９－８０Ｖ－及８２－１４０进行家兔体内感染，通过大体病变、组织病变和细胞病变的观察．研究其致病性及致病机理．结果表明：这两株Ｙ．ｅ．菌感染性与致病性明显弱于对照组──致病性血清型带毒力质粒Ｙ．ｅ菌株，其中．用４３９－８０Ｖ－菌株接种的家兔不发生感染与致病．用ＬＡＢ－Ｂ１８２菌株接种的家兔有排菌，肠粘膜上皮溃疡，急性脾炎病变．但无微脓肿及菌落形成．无组织细胞变性坏死．     

小肠结肠炎耶氏菌实验性感染家兔微观病变观察

应用透射电子显微镜观察致病性血清型带毒力质粒Ｙ．ｅ．Ｇ１５１及ＬＡＢ－Ｂ１８２菌株感染的家兔，其肠道及脾脏有微脓肿及菌落形成，在微脓肿形成的区域，组织细胞变性坏死严重．用ＬＡＢ—Ｂ１８２菌株感染的家兔，其肝脏可见局灶性凝固性坏死．在上述脏器无微脓肿及菌落形成的部位，肉眼观察正常，但在电子显微镜下．却呈现不同程度的组织细胞病变．细菌不在细胞内增殖．致病性血清型丢失毒力质粒的４３９—８０Ｖ－菌株感染的家兔，其肠道、肝脏及脾脏组织细胞无病理学改变．     

运动发酵单胞菌定点整合质粒的构建

运动发酵单胞菌(Zymomonas mobilis)是目前发现的乙醇发酵能力最强的微生物之一,被认为是进行大规模乙醇生产的潜在菌种,但其可利用的碳源范围窄.对其进行分子生物学基础研究和基因工程改造的工作早在上世纪八、九十年代就开始了,但基于其独特的生物学特性,至今遗传操作手段仍不成熟.以大肠杆菌(E.coli)pBR328为出发质粒,分段插入带Z.mobilis乳酸脱氢酶基因ldh、分别作为左右同源臂、1.5 kb大小的DNA片段,构建了Z.mo-bilis定点整合型质粒pBR328-ldhR-ldhL(6 239 bp),此质粒两同源臂片段即ldhR和ldhL的中间有稀有酶切位点NotI和SfiI,以方便插入待整合片段.经Notl位点插入1 216 bp氯霉素抗性基因cml,将相应质粒pBR328-ldhR-cml-ldhL以环型和线型形式分别电击转化Z.mobilis ZM4菌株,都成功筛选到了ZM4 ldh::cml菌株,证明此定点整合质粒是方便可用的,为对Z.mobilis的定点遗传操作提供了很好的基础.     

柳杉毛虫高毒力白僵菌Bbd3菌株的筛选及应用

对天然柳杉毛虫僵虫组织分离纯化,选取8株菌株在PDA 虫尸培养基上培养,在比较菌粉质量以及不同菌株室内外毒力测定试验基础上,筛选出菌丝生长快、产孢时间短、产孢量高、对柳杉毛虫毒力强的2株Bbd3和Bbd6高毒力白僵菌优良菌株,将具最优品质的Bbd3菌株大规模生产和大面积防治应用,菌粉平均含孢量达3.09×1010个/g,平均孢子萌发率达99.1%,平均防治效果达92%以上。     

一种酿酒酵母同源重组载体的筛选

将携有mTn-3xHA／LacZ转座子的酿酒酵母文库质粒pHSS6转化大肠杆菌DH5α,挑取单菌落并提取其质粒．将纯化质粒分别用Not Ⅰ酶切后,转化尿嘧啶缺陷型(ura^-)酿酒酵母菌株INVScl,使其同源重组到酿酒酵母基因组中,于SC／ura^-平板上筛选．取长有单菌落酵母的平板进一步筛选重组后上游有强启动子酿酒酵母文库质粒,用近300个质粒转化酵母菌株INVScl,最终得到2株前端有强启动子的酿酒酵母质粒,这2个质粒可作为酿酒酵母同源重组载体广泛应用．     

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.     

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.     

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.     

Characterization of Ordovician carbonate reservoirs, southeastern Saskatchewan, Canada

The discovery of the prolific Ordovician Red River reservoirs in 1995 in southeastern Saskatchewan was the catalyst for extensive exploration activity which resulted in the discovery of more than 15 new Red River pools. The best yields of Red River production to date have been from dolomite reservoirs. Understanding the processes of dolomitization is, therefore, crucial for the prediction of the connectivity, spatial distribution and heterogeneity of dolomite reservoirs.The Red River reservoirs in the Midale area consist of 3～4 thin dolomitized zones, with a total thickness of about 20 m, which occur at the top of the Yeoman Formation. Two types of replacement dolomite were recognized in the Red River reservoir: dolomitized burrow infills and dolomitized host matrix. The spatial distribution of dolomite suggests that burrowing organisms played an important role in facilitating the fluid flow in the backfilled sediments. This resulted in penecontemporaneous dolomitization of burrow infills by normal seawater. The dolomite in the host matrix is interpreted as having occurred at shallow burial by evaporitic seawater during precipitation of Lake Almar anhydrite that immediately overlies the Yeoman Formation. However, the low δ18O values of dolomited burrow infills (-5.9‰～ -7.8‰, PDB) and matrix dolomites (-6.6‰～ -8.1‰, avg. -7.4‰ PDB) compared to the estimated values for the late Ordovician marine dolomite could be attributed to modification and alteration of dolomite at higher temperatures during deeper burial, which could also be responsible for its 87Sr/86Sr ratios (0.7084～0.7088) that are higher than suggested for the late Ordovician seawaters (0.7078～0.7080). The trace amounts of saddle dolomite cement in the Red River carbonates are probably related to "cannibalization" of earlier replacement dolomite during the chemical compaction.     

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.     

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.     

橡胶颗粒沥青混合料低温抗裂性试验研究

以AC-13级配为基础,将橡胶颗粒代替部分集料掺入混合料中,以低温弯曲试验为评价方法对不同橡胶颗粒掺量下沥青混合料的低温抗裂性进行研究,并引入应变能密度值对混合料的低温抗裂性进行综合评价.试验结果表明:橡胶颗粒沥青混合料试件的破坏微应变均超过2 300,满足冬寒区的技术指标;无论是否掺加橡胶颗粒,随着温度的下降,沥青混合料破坏时的最大弯拉强度增大,弯拉应变降低,劲度模量增大;弯曲应变能密度在胶粒掺量为1%左右时具有较大的弯曲应变能密度值,此时橡胶颗粒沥青混合料具有较好的低温抗裂性.     

A computer generator for randomly layered structures

AcomputergeneratorforrandomlylayeredstructuresYUJia shun1,2,HEZhen hua2(1.TheInstituteofGeologicalandNuclearSciences,NewZealand;2.StateKeyLaboratoryofOilandGasReservoirGeologyandExploitation,ChengduUniversityofTechnology,China)Abstract:Analgorithmisintrod…     

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。     

Quantitative trait loci （QTLs） for quality traits related to protein and starch in wheat

Quality traits in wheat （Triticum aestirum L.） were studied by quantitative trait locus （QTL） analysis in a recombinant inbred line （RIL） population, a set of 131 lines derived from Chuan 35050 × Shannong 483 cross （ChSh）. Grains from RILs were assayed for 21 quality traits related to protein and starch. A total of 35 putative QTLs for 19 traits with a single QTL explaining 7.99-40.52% of phenotypic variations were detected on 10 chromosomes, 1D, 2A, 2D, 3B, 3D, 5A, 6A, 6B, 6D, and 7B. The additive effects of 30 QTLs were positive, contributed by Chuan 35050, the remaining 5 QTLs were negative with the additive effect contributed by Shannong 483. For protein traits, 15 QTLs were obtained and most of them were located on chromosomes 1 D, 3B and 6D, while 20 QTLs for starch traits were detected and most of them were located on chromosomes 3D, 6B and 7B. Only 7 QTLs for protein and starch traits were co-located in three regions on chromosomes 1D, 2A and 2D. These protein and starch trait QTLs showed a distinct distribution pattern in certain regions and chromosomes. Twenty-two QTLs were clustered in 6 regions of 5 chromosomes. Two QTL clusters for protein traits were located on chromosomes 1D and 3B, respectively, three clusters for starch traits on chromosomes 3D, 6B and 7B, and one cluster including protein and starch traits on chromosome 1D.