Cloning and Expression of the Leukotoxin BSBSE Gene from Fusobacterium necrophorum

以牛源坏死梭杆菌FNn株为试材,根据GenBank上已发表的坏死梭杆菌AF312861标准菌株的lktA序列设计1对引物,利用PCR技术扩增出1 131 bp的坏死梭杆菌白细胞毒素BSBSE基因。将PCR产物插入pGEM-T Easy vector中,经双酶切鉴定正确后进行序列测定。分析表明该BSBSE序列与GenBank上已发表的坏死梭杆菌AF312861标准菌株的lktA序列的核苷酸同源性为99%,推导出的氨基酸序列同源性为98%。为研究BSBSE的免疫原性,构建了原核表达载体pMAL-p2X-BSBSE,用IPTG诱导在大肠杆菌中表达。结果表明,BSBSE基因在大肠杆菌中进行了高效特异性融合表达,融合蛋白分子量约为84.5×103,其中41.5×103为BS-BSE基因表达的蛋白质,43.0×103为MBP融合标签,Western-blotting检测表明该表达产物有免疫原性。According to the sequence of announced lktA gene in Fusobacterium necrophorum,a pair of primers were designed.The BSBSE gene was amplified by PCR.The product was cloned into pGEM-T Easy vector.When nucleotide sequence and deduced amino acid sequence were compared with homologous sequence of the FN AF312861 lktA of GenBank,the homologue of the mucleotide sequence is 99% and the homologue of the amino acid sequence is 98%.The BSBSE fragment was inserted into expression vector pMAL-p2X and the plasmid pMAL-p2X-BSBSE were expressed in E.coli BL21 by IPTG induction.The SDS-PAGE analysis indicated the weight of the fusion protein was about 84.5.0×103,which included the 41.5×103 protein expressed from BSBSE gene and 43.0×103 fusion MBP tag.The recombinant BSBSE-pMAL-p2X production has Immunogenicity with western-blotting.The cloning and expression of the BSBSE gene established the foundation of further research on the function and application of the BSBSE gene.“十五”国家科技攻关子课题(2002BA518A04);; 中国农业科学院特产研究所科研基金项目(Tcs2005-03

伴有童年创伤的重症抑郁患者异常全脑功能连接：一项静息态功能磁共振研究

目的童年创伤（CT）被认为是成年后发展为重度抑郁（MDD）的主要危险因素之一。然而，伴有CT的MDD（CT-MDD）患者的神经基础仍然知之甚少。因此，本研究的目的是探讨CT-MDD患者静息态全脑功能连接（FC）。方法对34名CT-MDD患者和34名健康对照组进行静息态功能磁共振扫描。进行全脑体素水平度中心性（DC）分析，选取两组间显著差异的脑区作为感兴趣区（ROI），进一步计算全脑FC。随后，将异常脑区DC， FC值与临床量表进行相关性分析。结果CT-MDD组右侧额中回（MFG）的DC值高于健康对照组。基于种子点的脑功能连接分析发现，与健康对照组相比，CT-MDD组右侧MFG与右内侧前额叶和左侧楔前叶之间的FC增加（P<0.05）。此外，右侧MFG的DC值与CT严重程度相关。结论我们的结果表明，以右侧MFG作为ROI，其与默认网络（DMN）中的两个重要脑区右内侧前额叶和左侧楔前叶之间的FC增加，这可能表明CT-MDD患者的认知执行网络与DMN之间的同步性增加。这些发现可能为深入了解CT-MDD的病理生理机制提供了见解

体外循环中低流量肺动脉灌注对肺功能的影响

【目的】探讨体外循环中低流量肺动脉灌注对肺功能影响的作用机理。【方法】体质量18-22kg的杂种犬10只，随机分为对照组（n=5）和肺灌注组（n=5）。常规建立体外循环。肺灌注组用氧合血30mL／（kg·min）灌注肺动脉。分别于转流前，停体外循环即刻，转流后1h，转流后2h采集数据，计算氧合指数、肺动态顺应性。检测肺静脉血丙二醛（MDA）、超氧化物歧化酶（SOD）、细胞间黏附分子1（ICAM-1）和P选择素。【结果】转流后肺灌注组犬的肺动态顺应性和氧合指数明显高于对照组（P〈0．05）。两组肺静脉血MDA和SOD总体差异有统计学意义（P〈0．05），肺灌注组MDA含量低于对照组，SOD活性高于对照组。肺灌注组肺静脉血ICAM-1、P选择素含量均低于对照组（P〈0．05）。【结论】低流量持续肺动脉灌注通过减轻肺缺血再灌注损伤和粘附分子的激活来降低体外循环中的肺损伤

In Situ Raman Spectroelectrochemical Characterization of the Prussian Blue/Platinum Modified Electrode

采用现场拉曼光谱电化学技术表征了普鲁士蓝膜修饰铂电极的循环伏安过程 .结果显示 ,随着修饰膜的微观结构由普鲁士蓝向普鲁士白或相反过程转化 ,表征两种不同结构的拉曼特征振动谱峰及其强度变化呈现出明显的可逆特征 .Prussian Blue is known to be deposited electrochemically on the electrode surface, producing a dense redox active layer. Since the Prussian Blue modified electrode was first reported by Neff in 1978 [1] , different in situ spectroelectrochemical techniques have been widely used to investigate its electrochemical mechanism [2～4] . But no research work related to the in situ Raman spectroscopic studies on it has been reported. In this paper we try to characterize the nature of thin Prussian Blue films on platinum substrates with in situ Raman spectroelectrochemical technique. A Super LABRAM confocal microscopic Raman spectrometer (Dilor) with a He?Ne laser (632.8 nm) and CHI604A electrochemical Analyzer (USA) were used here. Cyclic voltammetry was used to prepare Prussian Blue film modified platinum electrode. The platinum electrode potential was cycled at a scan rate of 50 mV/s between 0 and 0.5 V (vs. SCE). As shown in figure1, the in situ Raman spectra obtained here characterized two cycles of voltammetric process of the Prussian Blue modified platinum electrode in a potassium chloride solution (1 mol/L) between 0.5 V and -0.1 V (vs.SCE) with a scan rate 1 mV/s. The observed spectra demonstrated the change of intensities and shifts of those characteristic Raman frequencies related to the coordination structures of Prussian Blue and its reduced form, Prussian White. From these spectra, intensity changes of those related characteristic Raman lines characterized the stability and reversibility of the Prussian Blue film obviously. As shown in Fig.2(a), at the beginning of the process, the characteristic Raman frequencies of Prussian Blue remarkably appeared at 2 089 cm -1 , 2 123 cm -1 and 2 154 cm -1 . When the scanning potential arrived at the range 0.20～0.15 V, which is somewhat lower than the reduction potential of Prussian Blue film modified platinum electrode, three peaks can be observed at 2 058 cm -1 , 2 109 cm -1 and 2 140 cm -1 respectively [Fig. 2(b)]. Finally, as shown in Fig. 2(c), at the potential range of -0.10～-0.05 V, characteristic peaks are located at about 2 048 cm -1 , 2 095 cm -1 and 2 130 cm -1 . At this moment, intensities of these peaks decreased to their minimum. These Raman frequencies should be related to the reduced state of Prussian Blue, i.e. Prussian White. Subsequently, with the potential scanning inversely, the intensities and shifts of these characteristic Raman lines changed reversibly.作者联系地址：上海师范大学化学系!上海200234,上海师范大学化学系!上海200234,上海师范大学化学系!上海200234,上海师范大学化学系!上海200234Author's Address: Dept. of Chem., Shanghai Teachers Univ., Shanghai 200234,Chin

Cloning and Expression of the Leukotoxin Gene SH from Fusobacterium necrophorum

坏死梭杆菌白细胞毒素是坏死杆菌病的主要致病因子,白细胞毒素基因(lkT)是其编码基因。以分离到的国内牛源坏死梭杆菌fn(A)菌株f4基因组dnA为模板,应用PCr方法扩增白细胞毒素基因SH片段,克隆至PMd18-T载体上,以bAMHⅠ和HIndⅢ酶切的目的片段SH与相应酶切的PET32A载体连接构建PET32A-SH重组表达质粒,经转化E COlI bl21(dE3)后用IPTg进行蛋白诱导,SdS-PAgE检测重组蛋白表达情况。结果表明:扩增基因序列大小为1800bP,SdS-PAgE检测重组蛋白有效表达,表达得到大小为80.2kdA的目的蛋白,采用镍柱亲和层析方法纯化SH重组蛋白,获得了纯度达95%的重组蛋白;经WEST-Ern-blOT证实,该蛋白对抗坏死杆菌阳性血清具有反应活性。The leukotoxin of Fusobacterium necrophorum(FN) is considered to be one of the main virulence factors.The lkt gene encodes for FN.In this study,the SH fragment of lkt gene was amplified by PCR using the F4 genome as the template,which was isolated from the Chinese Fusobacterium necrophorum strain.The fragment was then cloned to the pMD18-T vector for sequencing.Thereafter,the SH fragment was subcloned into the multiple cloning sites of the pET32 to construct pET32a-SH recombinant plasmid,which was then trans-formed into E.coli BL21(DE3) with IPTG induction for expression.SDS-PAGE was used to analyze the recombinant protein.The results showed that the SH fragment of about 1800 bp was amplified and was about 80.2 kDa.The fusion protein was purified by Ni-NTA affinity chromatography under denature conditions,and their purity was above 95%.Western-blot analysis indicated the SH fragment had anti-genicity against Fusobacterium necrophorum.“十五”国家科技攻关子课题(2002BA518A04);吉林省科技发展计划项目(20070570);吉林市科技发展计划项目(200805

the security design in a collaborative training system

文章根据一种协同训练系统中的特殊安全需求,分析了要解决的安全问题,并对用户登录验证、用户权限管理以及远程资源访问验证协议等方面提出了的设计方案以及部分实现

环境分析中生物监测技术的有效应用

保护环境、污染治理是当今社会发展最重要的环境课题。环境监测主要对各种环境污染因子进行监测,对制定环保政策及强化环境管理起着重要作用。加强环境保护管理必须以监测数据为基。若没有环境监测及环境监测质量不高,就无法对环境进行科学治理。</jats:p

Internet化学资源导航系统中的相关资源链接

化学家利用Internet通用资源搜索引擎如Yahoo、Google获取Internet资源时,检索结果常包含大量相关性较小的内容。Internet化学资源导航系统如ChemDex用人工方法收集和组织资源使资源内容质量和相关性比通用搜索引擎有所提高,但进行细致的分类仍比较困难。由中国科学院过程工程研究所建立、Internet化学化工资源导航系统ChIN的维护工具ChIN-Manag-er采用把两个内容相关的资源简介页互链来表示资源之间的密切相关关系,目前维护人员主要通用测览分类目录来确定相关资源,但这种方式在被索引的资源数量日益增大时其可用性降低。本论文开发了一种新的基于ChIN数据库检索的链接两个关系密切的相关简介页的方法。该方法针对ChIN数据库中简介页的组织特点设计了面向不同资源类型数据表的多种检索策略,这些策略侧重于被检索字段的确定;并为20余种不同类型的资源描述表建立了实现相应检索策略的检索界面。这些检索界面无缝集成到了ChIN-Manager相应的资源编辑界面中,为ChIN提供了一种快速确定被索引资源的密切相关资源的方法

Internet化学资源导航系统中的相关资源链接

化学家利用Internet通用资源搜索引擎如Yahoo、Google获取Internet资源时,检索结果常包含大量相关性较小的内容.Internet化学资源导航系统如ChemDex用人工方法收集和组织资源使资源内容质量和相关性比通用搜索引擎有所提高,但进行细致的分类仍比较困难.由中国科学院过程工程研究所建立、Internet化学化工资源导航系统ChIN的维护工具ChIN-Manager采用把两个内容相关的资源简介页互链来表示资源之间的密切相关关系,目前维护人员主要通用浏览分类目录来确定相关资源,但这种方式在被索引的资源数量日益增大时其可用性降低.本论文开发了一种新的基于ChIN数据库检索的链接两个关系密切的相关简介页的方法.该方法针对ChIN数据库中简介页的组织特点设计了面向不同资源类型数据表的多种检索策略,这些策略侧重于被检索字段的确定;并为20余种不同类型的资源描述表建立了实现相应检索策略的检索界面.这些检索界面无缝集成到了ChIN-Manager相应的资源编辑界面中,为ChIN提供了一种快速确定被索引资源的密切相关资源的方法