人RP2基因在果蝇胚胎细胞中的表达

将质粒pJLA503中的RP2编码cDNA用PCR的方法扩增,并与pIND-3myc质粒上的人c-myc编码序列连接到果蝇转基因载体pUAST上,构建了UAS－RP2－3myc和UAS－RP2(del6)-3myc融合质粒。用脂质体转染的方法,将质粒导入果蝇培养细胞系Schneider line2(S2),并用PCR加以证实,然后通过Western blot检测证明了RP2和RP2 del6的表达。证明了人的RP2基因在果蝇细胞中表达的可能性,说明果蝇系统是可以用来分析人的RP2基因功能的。     

dRaf1转基因果蝇的构建与表型分析

黑腹果蝇(Drosophila melanogaster)是生物学研究中最重要的模式生物之一.首先,将dRaf1基因克隆到Gal4诱导表达的染色体定点插入载体p UAST-att B质粒后进行显微注射,成功构建了可诱导dRaf1表达的转基因果蝇.将dRaf1转基因果蝇品系与眼睛或翅膀特异性表达GMR-Gal4和Nub-Gal4品系果蝇分别杂交,观察dRaf1在果蝇或翅膀过表达的表型,确定其发育特征.其次,将翅膀中过表达dRaf1的果蝇与HpoRNAi的果蝇杂交发现,dRaf1在果蝇翅膀中的表型明显增强.本研究为大规模筛选修饰dRaf1活性与功能的调节因子奠定基础.     

果蝇锌离子转运蛋白Catsup亚细胞定位分析

Catsup是果蝇体内一个功能重要的基因,其突变会导致体内多巴胺含量过高,引起果蝇致死及生殖障碍.绿色荧光蛋白(green fluorescent protein,GFP)在确定蛋白的亚细胞定位进而在研究蛋白功能方面起着重要的作用.为了探索Catsup的亚细胞定位进而能够更好地研究Catsup的功能,文章利用基因工程技术和生物学方法,聚合酶链式反应(polymerase chain reaction,PCR)扩增Catsup基因全长,与GFP一起连接至pUAST质粒,构建载体pUAST-Catsup-GFP,通过显微注射技术导入果蝇,获得UAS-Catsup-GFP转基因果蝇.通过克隆验证了转基因果蝇构建成功.利用该果蝇进行Catsup的亚细胞定位,确定其定位在高尔基体上,为进一步研究该基因的功能奠定基础.     

EGFP和ALR融合基因表达载体构建及其在HeLa细胞中的表达

从人血液中提取总DNA，利用PCR技术扩增人肝细胞再生增生因子基因，将其插入表达载体pEGFP—C1的多克隆位点中，构建增强绿色荧光蛋白（enhanced green fluorescence protein gene，EGFP）和人肝细胞再生增强因子（human augmenter of liver regeneration gene，ALR）融合基因表达载体pEGFP／ALR，并将其转染Hela细胞系，用含G418的DMEM／F12培养液筛选转基因细胞，然后利用PCR和聚丙烯酰胺凝胶电泳检测转基因细胞中ALR基因的存在及其表达，用荧光显微镜检测EGFP基因的表达．结果显示：得到了构建正确的EGFP和ALR融合基因表达载体；在转基因细胞中，PCR扩增得到1．7Kb的ALR条带，蛋白电泳得到57KD大小条带．与ALR和EGFP融合蛋白大小相符；荧光显微镜下观察到发绿色荧光的Hela细胞．在转基因细胞中，EGFP和ALR同时存在并表达，绿色荧光蛋白可作为报告基因指示目的基因的表达，从而简化了目的基因繁琐的检测手段．     

pEGFP-N3-t-PA真核表达质粒的构建及其在成纤维细胞中的表达

目的利用基因工程技术克隆人组织纤溶酶原激活物（t—PA）基因并构建一种能高效、安全表达t—PA的pEGFP—N3-t—PA真核表达重组质粒,为进一步研制转基因动物奠定基础．方法采用高效Trizol试剂快速从黑色素瘤细胞中提取总RNA,RT—PCR获得t—PAcDNA,并将真核表达质粒pEGFP—N3和t—PA基因片段分别双酶切,将回收的pEGFP—N3大片段（4．7kb）与t—PA基因片段（1．1kb）重组．对pEGFP—N3-t—PA质粒进行酶切鉴定和基因测序鉴定．脂质体介导pEGFP—N3-t—PA转染成纤维细胞（NIn313）,并用RT—PCR法从mRNA水平检测t—PA的表达情况,用倒置荧光显微镜检测、分析其在NH 3T3细胞中的表达．结果成功地从黑色素瘤细胞中克隆了t—PA基因,并构建了以pEGFP—N3为载体的真核表达质粒载体,并能在真核细胞中表达、分泌t—PA．结论含t—PA基因真核表达质粒构建成功．     

dDnmt2与dMBD2/3在果蝇不同发育时期的转录表达

为了深入了解果蝇DNA甲基化修饰系统及其相关基因的表达特点及功能,以黑腹果蝇Canton S品系为材料,应用实时荧光定量PCR方法检测分析不同发育时期果蝇dDnmt2与dMBD2/3基因的转录表达.结果表明:dDnmt2与dMBD2/3分别在6～9 h和0-3 h胚胎期表达量最高,在12～15 h和18～21 h胚胎、三龄幼虫、蛹、成虫期表达明显降低.果蝇dDnmt2和dMBD2/3基因的表达特点与果蝇不同发育时期基因组DNA甲基化水平相一致.     

通过遗传转化获得含多基因的水稻转基因纯合植株

利用基因枪法将含有4个不同基因的3个质粒共转化由粳稻品种鄂宜105号和鄂晚5号种子胚诱导的愈伤组织（5－10d龄）。从轰击的986块愈合组织中共再生出169株独立的转基因水稻植株（转化率为17％）。PCR／Southern blot分析显示70％以上的转基因植株含有所有4个基因。GUS组织化学分析、Western blot和或RT－PCR分析表明所有4个基因的共表达率为70％。未观察到任何质粒在整合中存在优势,转基因拷贝数也与基因表达量无关。遗传分析证实外源基因在后代植株中大多以孟德尔方式遗传。从其R1代为3：1孟德尔方式遗传的后代R2代植株中,鉴定含有3个或4个不同基因的转基因纯合植株系。PCR／Southern blot分析证实了这些转基因纯合植株系。这些系的植株具有相似的外源基因表达量。我们证实通过基因枪介导的共转化,结合常规育种方法筛选可以获得含多基因的转基因水稻纯合植株。这项技术为利用基因同时改良作用多个性状提供了一种途径。     

利用目的基因转化技术培育棉花抗病新品种

用花粉管通道法将菜豆几丁质酶基因与烟草β－1、3葡聚糖酶基因双价植物表达载体pBLGC导入新疆棉花主栽品种（系）系550、822、1304、石远321。通过卡那霉素检测、PCR－Southern验证、抗黄萎病、枯萎病性筛选，得到抗病性良好且遗传性稳定的转基因棉花。经过6年10个世代的选育，育成抗枯萎病、抗（耐）黄萎病的转基因抗病新品系。     

转基因花卉的DNA提取与多重PCR分析

分别以CTAB法、DNA提取试剂盒提取菊花、矮牵牛、非洲菊3种花卉的基因组DNA，凝胶电泳结果表明试剂盒提取的效果较好。设计合成了3对引物，分别扩增花椰菜花叶病毒（Cauliflower mosaic virus,CaMV)35S启动子、根癌农杆菌胭脂碱合成酶基因（nos）终止子、大肠杆菌K12菌株新霉素磷酸转移酶Ⅱ(nptⅡ）基因。普通PCR检测结果表明，3种花卉6个样品中仅矮牵牛的1个样品含有这3种转基因成分，酶切鉴定结果验证了PCR产物为非假阳性。尝试以多重PCR同时检测转基因矮牵牛与对照阳性质粒中的2个或3个转基因成分，得到预期结果。     

抗汉滩病毒单抗3G1 scFv植物表达载体的构建

利用PCR方法，从含有3G1 scFv基因的重组质粒中扩增出抗体基因，并使基因两端携带合适的限制性酶切位点．将其克隆人植物表达载体pBI121，构建获得3G1 scFv-pBI121重组质粒．酶切鉴定及测序结果均证明重组质粒构建成功，将重组植物表达载体转入农杆菌LBA4404，为进一步构建转基因植物的研究奠定了基础．     

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.     

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.     

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.     

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.     

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.     

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.     

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.