新城疫病毒La Sota疫苗株基因组全长cDNA克隆的构建及鉴定

为构建能为新城疫病毒拯救提供直接使用的全长cDNA克隆,利用分子克隆的方法,将新城疫病毒La Sota株全基因组分成末端部分重叠的11个片段(F1～F11),按其基因组顺序克隆至改造后的真核表达载体pVAXr上,同时在全长cDNA两侧引入T7启动子和丁型肝炎病毒核酶(Rib)序列,以确保病毒基因组转录产物的两端形成精确的核酸序列。鉴定结果表明,新城疫病毒La Sota株基因组全长cDNA已成功克隆至pVAXr上,命名为pVAXr-FL。本研究为新城疫病毒的拯救及外源基因的表达奠定了基础。     

骆驼斯氏副柔线虫病传播媒介的筛选与确定

对从内蒙古骆驼生活环境中采集到的24种蝇进行普通生物学剖检,并进一步用斯氏副柔线虫特异性引物ST1、ST2对蝇体内检获的幼虫进行分子生物学鉴定,以筛选与确定骆驼斯氏副柔线虫病的传播媒介。结果显示,在西方角蝇和截脉角蝇体内均发现了疑似斯氏副柔线虫幼虫,感染强度为1～22条/个,雌蝇与雄蝇感染强度差异不显著;幼虫长度为1.033～1.934mm;雄蝇的感染率为43.8%～48.0%,雌蝇的感染率为35.7%～46.2%。扩增得到的部分ITS序列(包括5.8S)与GenBank中登录的斯氏副柔线虫ITS序列同源性高达99.3%。基本确定了西方角蝇和截脉角蝇为斯氏副柔线虫病的传播媒介。     

鸡贫血病毒VP1、VP2和VP3基因酵母双杂交载体的构建及其自激活活性的鉴定

为构建鸡贫血病毒(CAV)VP1、VP2、VP3基因酵母双杂交载体并鉴定其自激活作用,从CAV细胞传代毒M9905毒株中提取基因组DNA,利用PCR分别扩增VP1、VP2和VP3基因,采用Gateway技术将VP1、VP2和VP3基因分别克隆到酵母双杂交载体pDEST32和pDEST22中,构建了诱饵载体及猎物载体,经酶切和PCR鉴定且测序正确后,用醋酸锂法转化酵母菌株Mav203,通过缺陷型平板SD/-Leu/-Trp/-His和SD/-Leu/-Trp/-Ura筛选以及LacZ报告基因检测载体有无自激活作用。结果表明,成功构建了诱饵载体pDEST32-VP1/VP2/VP3和捕获载体pDEST22-VP1/VP2/VP3,并证明其VP1、VP2和VP3蛋白在酵母双杂交系统中无自激活作用。研究结果为下一步利用酵母双杂交系统探索CAVVP1、VP2、VP3蛋白之间的相互作用奠定了基础。     

中药富硒酵母对闽中麻鸡肝和小肠细微结构的影响

选择1日龄闽中麻鸡396只,随机分为4组,A组饲喂玉米-豆粕型基础日粮,B组、C组、D组在基础日粮中分别添加2、3、4 g/kg的中药富硒酵母,试验为期49 d,研究了日粮中添加中药富硒酵母对闽中麻鸡肝和小肠发育的影响.于第7、21、35、49 d每组随机抽取6只鸡,采取肝和小肠各段,Bouin液固定,制作石蜡切片,光镜观察;第49 d取A组和D组的肝和空肠,25 mL/L戊二醛固定,制作超薄切片,电镜观察.结果显示,与对照组相比,试验组肝的平均质量和器官指数大多高于对照组;肝细胞线粒体增多增大,枯否氏细胞内溶酶体增多;小肠绒毛和肠腺长于A组;空肠柱状细胞的微绒毛增长.证实,基础日粮中添加中药富硒酵母能促进鸡肝和小肠的发育,改善肝细胞和小肠黏膜的细微结构.     

pIRES2-EGFP-LR真核表达载体的构建及其在293T细胞中的表达

为探讨层黏蛋白受体的生理功能及其在疾病发生、发展过程中的作用,以培养的SD大鼠大脑皮质神经元的总RNA为模板,运用RT-PCR方法扩增了SD大鼠层黏蛋白受体基因.将其克隆到pMD18-T载体中,经酶切纯化后再将其亚克隆到带有加强型绿色荧光蛋白报告基因的pIRES2-EGFP真核表达栽体中,对重组质粒pIRES2-EGFP-LR进行酶切和测序鉴定后,采用阳离子脂质体转染法将重组质粒转染到293T细胞,荧光显微镜和激光共聚焦显微镜观察其表达情况.结果表明,真核表达载体plRES2-EGFP-LR构建成功并可以在293T细胞中表达.该表达载体可以作为研究层黏蛋白受体生理功能的重要工具.     

口蹄疫病毒多基因腺病毒穿梭质粒的构建

采用分步扩增连接的方法将口蹄疫病毒结构蛋白基因P1 2A、3C蛋白酶基因和 3D聚合酶基因按正确的读码框依次连接克隆入 pGEM T载体。然后 ,将切取的目的基因亚克隆入腺病毒穿梭质粒 pShuttle CMV。构建成功的 2个重组穿梭质粒经测序鉴定 ,含有完整的目的基因表达盒 ,穿梭质粒可与腺病毒骨架载体进行细菌内同源重组 ,以产生重组腺病毒载体     

猪瘟病毒E2基因主要抗原区原核表达载体的构建及表达

用RT PCR技术对猪瘟兔化弱毒C株、石门强毒株 (Shimen)、近期地方流行的属于第 1群毒株的新疆毒株 (XJ)和第 2群代表毒株甘肃临洮 (LT)株E2基因的主要抗原区进行扩增 ,均得到约 5 6 1bp的基因片段 ;经克隆测序及序列分析 ,该基因编码 187个氨基酸残基 ,预计蛋白分子质量为 2 1.7ku。将这 4个基因分别克隆到原核表达载体 pGEX 4T 1中 ,经酶切、PCR扩增和测序分析确证其正确插入到表达载体中 ,阅读框是正确的 ,从而构建成重组表达载体。重组质粒转化至宿主菌BL2 1(DE3)中 ,用异丙基硫代 β D 半乳糖苷 (IPTG )进行诱导表达 ,对表达的蛋白用SDS PAGE电泳、免疫印迹和薄层凝胶扫描分析。结果表明 ,E2基因的主要抗原区可以在大肠埃希氏菌中表达 ,表达的蛋白多以包涵体形式存在 ,表达产物的分子质量约为 5 0 .0ku ,与理论推测的分子质量一致 ;薄层凝胶扫描分析表明 ,表达蛋白约占菌体蛋白总量的 2 0 .1% ;免疫印迹结果证明 ,表达的E2蛋白可被猪瘟阳性血清所识别。     

Automated Classification of Epiphyses in the Distal Radius and Ulna using a Support Vector Machine

The aim of this study was to automatically classify epiphyses in the distal radius and ulna using a support vector machine (SVM) and to examine the accuracy of the epiphyseal growth grades generated by the support vector machine. X‐ray images of distal radii and ulnae were collected from 140 Chinese teenagers aged between 11.0 and 19.0 years. Epiphyseal growth of the two elements was classified into five grades. Features of each element were extracted using a histogram of oriented gradient (HOG), and models were established using support vector classification (SVC). The prediction results and the validity of the models were evaluated with a cross‐validation test and independent test for accuracy (P_A). Our findings suggest that this new technique for epiphyseal classification was successful and that an automated technique using an SVM is reliable and feasible, with a relative high accuracy for the models.     

Classification of firing pin impressions using HOG-SVM

Crimes, such as robbery and murder, often involve firearms. In order to assist with the investigation into the crime, firearm examiners are asked to determine whether cartridge cases found at a crime scene had been fired from a suspect's firearm. This examination is based on a comparison of the marks left on the surfaces of cartridge cases. Firing pin impressions can be one of the most commonly used of these marks. In this study, a total of nine Ruger model 10/22 semiautomatic rifles were used. Fifty cartridges were fired from each rifle. The cartridge cases were collected, and each firing pin impression was then cast and photographed using a comparison microscope. In this paper, we will describe how one may use a computer vision algorithm, the Histogram of Orientated Gradient (HOG), and a machine learning method, Support Vector Machines (SVMs), to classify images of firing pin impressions. Our method achieved a reasonably high accuracy at 93%. This can be used to associate a firearm with a cartridge case recovered from a scene. We also compared our method with other feature extraction algorithms. The comparison results showed that the HOG-SVM method had the highest performance in this classification task.     

The Dynamics of Mass Killings: Testing Time-Series Models of One-Sided Violence in the Bosnian Civil War

Many observers contend that wartime civilian victimization is an instrument of political leaders to achieve a particular goal. This article examines whether retaliation for similar acts by the other side, the developments on the battlefield, or the behavior of international actors accounts for the ups and downs of this so-called one-sided violence. Using information from the Konstanz One-Sided Violence Event Dataset and other sources, we evaluate the empirical relevance of these complementary models statistically. Time series analyses of the weekly number of killed and harmed Muslims (Bosniacs) and Serbs during the Bosnian civil war support the military and the massacre logic. We show that the Serbian side decreased one-sided violence following a territorial conquest, but that its one-sided violence was not a reciprocal response to the Bosniac targeting of civilians. Conversely, the Bosniac side resorted to violence during times of increasing Serbian atrocities and when the fighting was particularly intense. The analysis reveals that most international interventions did not reduce the carnage, but that the Serbs responded to Russian moves.