SARS—CoV N蛋白与病毒mRNA相互作用的初步研究

SARS冠状病毒(SARS-CoV)是一种新型的冠状病毒,其基因组大小约为30,000 nt,为单股正链RNA。病毒基因中的1-72个核甘酸为前导序列。核衣壳(Nucleocapsid,N)蛋白是冠状病毒的主要结构蛋白,它在病毒基因转录,翻译以及病毒颗粒包装中起重要作用。在本研究中,我们通过PCR的方法从SARS-CoV cDNA中克隆N基因,将基因克隆到大肠杆菌表达载体中,经表达纯化获得大量重组蛋白,通过亲和层析和凝胶过滤层析获得高纯度的N蛋白。同时构建前导RNA的转录模板,经体外转录得到地高辛标记的RNA。使用Northwestern分析技术,我们证实纯化的N蛋白在体外可以与RNA发生特异性的结合。N蛋白与病毒RNA的结合特性及其在病毒生活周期中的所起作用的初步研究,为下一步设计出有效的阻断病毒周期从而达到抗病毒目的的药物或疫苗奠定了基础。     

不同防治措施对刚竹毒蛾种群变动的影响

应用灰色关联度（the gray sequence）分析不同防治措施对刚竹毒蛾（Pantana phytlostachysae Chao）种群变动的影响。分析结果表明：生物防治标准地种群变动与对照区相似,虫口较低;化学防治后标准地虫口较对照区为高,种群变动与对照区差异较大。第1代和第3代中各防治区与对照区种群变动最接近的均是白僵菌粉炮,其次是敌敌畏烟剂和油烟剂。第2代则是白僵菌粉炮〉油烟剂〉敌敌畏烟剂。进一步说明不同防治措施在不同环境条件下,干扰自然生态系统的程度不同。     

小花棘豆(Oxytripis glabra DC)毒素基因的cDNA克隆及序列测定

在完成小花棘豆毒素 95 %氨基酸序列的基础上 ,根椐已知的氨基酸序列 ,设计合成了特异简并引物 .以小花棘豆总RNA为模板 ,逆转录合成cDNA第一链 ,用置换法合成双链cDNA .用特异引物对此双链cDNA进行PCR扩增 ,将扩增后的目的基因与用SmaⅠ酶切的质粒pUC 18连接 ,转化大肠杆菌JM10 7.筛选阳性克隆进行序列分析 ,获得了OXY基因的全部序列 .经测序后测得基因序列与原氨基酸序列对照完全一致 .GenBnak数据检索说明 ,OXY基因编码序列确定是一个从未报道的序列 .此研究结果对该毒素的应用研究奠定了基础 .     

对虾白斑杆状病毒DNA聚合酶调控序列的克隆

为了揭示对虾白斑杆状病毒的致病机理,将该病毒基因组中推测的DNA聚合酶上游调控序列克隆进荧光素酶报告基因载体中,以便寻找一个能表达该病毒基因的细胞系统.     

大头蛙Sox基因的克隆与序列分析

参考人SRY基因HMC—box的保守区序列,设计一对特异引物,采用PCR技术扩增了大头蛙的Sox基因,并对扩增产物进行了克隆与测序。结果在雌雄个体中均扩增出217bp的基因片段,与人对照相同。序列分析表明,大头蛙雌雄个体之间Sox序列没有差异,与人SOX基因的同源性达到88％,与其它各类动物的Sox基因也都有非常高的相似性。结果支持Sox基因在进化上十分保守的结论。     

早期形态发生:一种解决贻贝科(软体动物门:双壳纲)分类、系统发育和进化问题的方法

作者对贻贝科贝类的幼虫和幼贝期发育阶段形态结构的出现和变化顺序进行了研究,其约60个不同分类单元的个体发生可归纳为4种形态发生类型或模式。主要对3个形态发生区域的阶段形态结构的起源、发育变化和同源性做了研究。其一,即中央区域,开始形成于前双壳Ⅰ期(PD-Ⅰ),在某个分类单元它可以在前双壳Ⅱ期(PD-Ⅱ)和幼贝期(N)形成,而在其它分类单元则在前双壳Ⅱ期、幼贝期和双壳期(D)形成;第二区域,即背部后区,在幼贝期出现;第三区域,即背部前区,出现于双壳期。双壳期背部后区在某个分类单元起源于幼贝期的形态构造,在其它分类单元则可能起源于双壳期的形态构造。与在贻贝分类学上应用的成体特征相比,早期发育阶段中央和背部后区的形态结构显示出很明显的发育顺序或特征变化规律。根据以前人们熟知而尚未应用到分类和系统发育研究中的早期发育阶段形态特征,作者重新修订了Soot-Ryen的现生贻贝科种上阶元分类系统,重新提出了科内系统发育关系。修订的分类系统表明,Scarlato and Starobogatov(1984)提出的贻贝科各亚科由偏顶蛤亚科开始,沿4条系统发育路线演化发展,对应其早期发育阶段的4类形态发生类型或模式。     

阴道毛滴虫氢化酶体腺苷酸激酶基因的克隆及序列分析

目的-克隆阴道毛滴虫氢化酶体腺苷酸激酶(AK)基因,并测定其序列,进行序列分析。方法-根据AK基因已知序列设计合成一对引物,应用PCR技术从阴道毛滴虫基因组DNA中扩增出AK基因,并将其克隆入pMD18-T simple载体。阳性克隆的重组质粒经酶切及PCR鉴定后,用双脱氧链末端终止法进行基因序列测定。应用BLAST软件辅助分析所测基因与Genbank中阴道毛滴虫氢化酶体AK序列的同源性。结果-PCR扩增得到特异的阴道毛滴虫氢化酶体腺苷酸激酶基因序列。酶切及PCR鉴定获得了正确的PT-AK重组质粒。测序表明,所克隆的AK基因大小为690bp,编码229个氨基酸。序列分析表明,所测基因与Genbank中阴道毛滴虫氢化酶体AK序列具有高度同源性(99．9％)。结论-克隆了阴道毛滴虫氢化酶体腺苷酸激酶基因,序列测定及同源性分析表明,所测基因与Genbank中阴道毛滴虫氢化酶体AK序列具有高度同源性。     

Polymorphic sequence‐characterized codominant loci in the chestnut blight fungus,Cryphonectria parasitica

Studies on the population biology of the chestnut blight fungus, Cryphonectria parasitica, have previously been carried out with dominant restriction fragment length polymorphism (RFLP) fingerprinting markers. In this study, we describe the development of 11 codominant markers from randomly amplified polymorphic DNAs (RAPDs). RAPD fragments were cloned and sequenced, and polymerase chain reaction (PCR) primers were designed flanking insertions/deletions. Primers labelled with fluorescent dyes were combined in multiplex reactions to assay five or six loci simultaneously in a capillary sequencing system. These codominant markers have the potential to complement RFLP methods for studying C. parasitica.     

Isolation and characterization of microsatellite markers from the entomopathogenic fungus Metarhizium anisopliae

Fourteen polymorphic microsatellite markers were isolated from the entomopathogenic fungus, Metarhizium anisopliae, based on enriched genomic libraries. In order to assess allelic variability, the microsatellite loci were analysed in a collection of 34 isolates sampled from across Switzerland. The number of detected alleles in 14 loci ranged from two to eight and expected heterozygosity from 0.265 to 0.808. Because of the high expected heterozygosity, the 14 microsatellite loci are very useful for ecological studies and analysis of population diversity, and to identifying, monitoring, and tracking M. anisopliae strains applied as biological control agents.     

Development of polymorphic markers for Cirsium arvense,Canada thistle,and their amplification in closely related taxa

Suppression of invasive Canada thistle, Cirsium arvense, with biological control agents has stalled because introduced agents were not host‐specific. To aid in the development of more effective management strategies, molecular markers are needed to examine the genetic structure of Canada thistle populations. Microsatellite (simple sequence repeat) markers were developed and intersimple sequence repeat (ISSR) markers were tested for North American populations. An average of nine polymorphic alleles per microsatellite locus and 11 per ISSR locus were detected. These will be used to examine the genetic structure of C. arvense in the northern Great Plains and their transferability to endemic Cirsium spp.