His-猪生长激素融合基因在Bm-N细胞和蚕体内的表达与纯化

用构建的带有His—Tag pgh融合基因的重组杆状病毒Bm—BacPAK6—pgh研究了Bm—BacPAK6—pgh在家蚕细胞(Bm—N)和蚕体内的表达，并对蚕体表达产物进行了纯化．SDS—PAGE电泳分析显示，重组病毒Bin—BacPAK6—pgh在Bin—N细胞、蚕体中得到了融合表达，Western blot分析表明，Bm—N细胞、蚕体中表达的融合蛋白与大肠杆菌表达的猪生长激素具有相同的抗原性．Bm-BacPAK6-pgh在Bin—N细胞内的表达始于24h，而蚕体中的表达始于72h，二者的表达峰分别在96h和120h．经40％饱和度硫酸铵盐析和Ni—NTA Agarose亲和柱二步纯化可获得SDS—PA(正电泳纯的具有抗原性的重组猪生长激素融合蛋白。     

A型口蹄疫病毒（FMDV）VP1基因遗传进化分析

对疑似感染FMDV的样本进行了FMDV VP1基因RT-PCR鉴定,测定了其中1个阳性样本序列(命名为HY).采用生物信息学软件比较了HY与参考序列的同源性,并构建VP1基因的分子系统树.结果表明:HY属A型FMDV亚洲拓扑型东南亚-97谱系,与我国近年来报道的A/GDMM/CHA/2013的序列相似性为99.1%,与AF/72疫苗株的相似性仅为79.0%.提示我国近年来报道的A型毒株与AF/72疫苗株的序列差异较大,应及时更换A型口蹄疫疫苗毒株.     

中度嗜盐菌GbsB基因的克隆、序列分析及生物信息学分析

为构建耐盐的转基因植物提供材料,根据地衣芽孢杆菌Bacillus licheniformisATCC 14580的GbsB基因的核甘酸序列设计1对特异性引物,通过PCR的方法扩增由分离的中度嗜盐菌Bacillus sp.XJ1-05的GbsB基因,经T/A克隆,插入到pBS-T载体上进行序列测序,采用NCBI-BlastX软件在Genbank数据库中进行同源性检索,结果表明:得到的GbsB基因的开放阅读框(ORF)全长为1209bp,编码1个由402个氨基酸残基组成的蛋白质;其与Bacillus licheni-formisATCC 14580的GbsB基因的氨基酸序列同源性高达96%。生物信息学分析表明:该GbsB基因编码蛋白无信号肽,无跨膜区域,高亲水性,有2个乙醇脱氢酶作用位点。     

稠油降解菌的筛选及其生物表面活性剂的特性

添加稠油对土壤中土著微生物进行驯化, 分离出33株能以稠油为惟一碳源生长的细菌, 从中筛选出2株高效表面活性剂产生菌XJ1和SJ4, 9株高效稠油降解菌. XJ1和SJ4可将发酵液的表面张力由72.4 mN/m分别降到36.1 mN/m和36.2 mN/m;14 d摇瓶油降解率分别为35.89%和31.59%, 降解效率在各单菌中最高. 同时研究了发酵液中XJ1和SJ4的生长量与其生物表面活性剂产生情况之间的关系, 经红外光谱分析初步确定两种生物表面活性剂均为糖脂类化合物.     

Generation of VP5 deficient mutant of infectious bursal disease virus strain HZ2

Infectious bursal disease virus (IBDV) is one of the most significant viral pathogens in chickens[1]. It causes high mortality in young chickens and establishes an immunosuppression state by destroying the precursorsInfectious bursal disease virus (IBDV) …     

Evolution and molecular epidemiology of foot-and-mouth disease virus in China

Foot-and-mouth disease (FMD),caused by foot-and-mouth disease virus (FMDV),is considered one of the most important viral diseases of cloven-hoofed animals,causing severe economic losses in affected regions of the world.Three serotypes (A,O,and Asia 1) of FMDV have been identified in China since 1958.In addition,the occurrence of novel subtypes within these serotypes has made the epidemiology of FMDV more complicated over the last few years.In this review,we summarize the history and the current epidemiological situation in China,genetic diversity (e.g.,quasispecies dynamics,antigenic heterogeneity,and functional constraints),intertypic recombination,and the evidence for positive selection of different FMDV serotypes.We also assess these genetic data to understand the origin,evolution,and transmission of FMDV,the findings of which may be useful in developing control measures for future epidemics.     

Infectious foot-and-mouth disease virus derived from a cloned full-length cDNA of OH／CHA／99

The China foot-and-mouth virus (FMDV) isolate OH/CHA/99 was isolated from swine, which was unable to infect bovine thyroid cells in vitro or to cause typical disease in bovines following intradermal inoculation in the tongue. To enhance antigenicity, replication, maturation and pathogenicity studies of OH/CHA/99, an infectious fulllength cDNA clone, designated pBIFMDV, was prepared. The in vitro and in vivo biological properties of the virus derived from pBIFMDV were studied by analyzing antigenicity, plaque morphology and virulence in pigs. The results showed that the virus derived from pBIFMDV had the same biological properties as the parent strain OH/CHA/99; the fulllength infections cDNA clone, pBIFMDV, will be very useful in studies of the antigenicity, virulence, pathogenesis, maturation and replication of FMDV.     

Sequence analysis of a 7.3 kb Bam H Ⅰ genomic fragment of fowlpox virus strain 282E_4

A 7.3 kb BamH I genomic fragment from fowlpox virus (FPV) strain 282E4 has been sequenced. After comparison with the FPV sequences collected in Genbank, and the entire genomic sequence of vaccinia virus (VV) strain Compenhagen, a 11.2 kb BamH I fragment from FPV strain Munich HP-438 has been found homologous to the fragment. Every predicted open reading frame (ORF) within the fragment has been compared with the ORFs of W strain Copenhagen at the amino acid level. The signal peptide and transmembrane region of each peptide encoded by each corresponding ORF predicted in the fragment have been studied as well.     

Sequence analysis of a 7.3 kbBam H I genomic fragment of fowlpox virus strain 282E4

A 7.3 kb BamH I genomic fragment from fowlpox virus (FPV) strain 282E₄ has been sequenced. After comparison with the FPV sequences collected in Genbank, and the entire genomic sequence of vaccinia virus (VV) strain Compenhagen, a ll.2 kb BamH I fragment from FPV strain Munich HP-438 has been found homologous to the fragment. Every predicted open reading frame (ORF) within the fragment has been compared with the ORFs of VV strain Copenhagen at the amino acid level. The signal peptide and transmembrane region of each peptide encoded by each corresponding ORF predicted in the fragment have been studied as well.     

猪圆环病毒2型ORF2基因及片段的原核表达

目的表达猪圆环病毒2型ORF2抗原,为研制猪圆环病毒2型血清学诊断方法以及生物制品生物安全性检验,提供技术支持。方法根据猪圆环病毒2型(PCV2)LC株序列,设计合成5对引物,采用PCR方法从全序列重组质粒PCV2-LC中扩增出了ORF2基因和4个不同ORF2基因片段,分别将其克隆到原核表达载体PET-32a上,再分别将各个重组表达质粒转化到大肠杆菌BL21中,用终浓度1 mmol/LIPTG诱导。结果表达产物以包涵体的形式存在,经Western-blot检测表明,表达的重组蛋白ORF2b、ORF2c、ORF2d能够被PCV2阳性血清所识别,具有良好的抗原性。表达的重组蛋白为ELISA诊断试剂盒的研制奠定了基础。     

亚洲一型口蹄疫病毒YNAs1.1株结构蛋白VP1基因的核苷酸序列分析

提取BHK21细胞增殖的亚洲一型口蹄疫病毒（foot-and-mouth disease virus Serotype Asial)强毒株YNAs1.1的RNA,用一对引物P7,P13经反转录（RT）－PCR法扩增了约674bp的DNA片段。克隆目的基因后,采用双脱氧DNA链末端终止法测得了YNAs1.1的VP1基因36－633核苷酸序列。分析表明,病毒VP1基因的核苷酸序列与以色列以及印度已报道的Asia1型FMDV的同源性分别为82.11%与88.07%,对应的氨基酸序列同源性为87.94％与93.47％。该序列在GeneBank登陆号为AF241566。     

新基因Splt137在大肠杆菌中的表达及抗体制备

用PCR的方法扩增得到斜纹夜蛾核多角体病毒 (SpltMNPV)ORF137全长基因 (Splt137)。将其克隆到原核表达载体pQE30上 ,并转化大肠杆菌M15 ,在IPTG诱导下表达了与预期相对分子质量相符的一个 2 7× 10 7的蛋白质 ,经过聚丙烯酰氨凝胶电泳纯化 ,免疫家兔制备了抗Splt137抗体。Western免疫印迹分析表明 ,该抗体适合用作Splt137蛋白的进一步分析。     

猪Akt1、Akt2基因克隆与组织表达图谱分析

为了得到长白猪蛋白激酶Akt1和Akt2基因序列并分析其表达模式,本研究使用RT-PCR方法,首先克隆了蛋白激酶Akt1和Akt2的cDNA.序列分析显示:长白猪Akt1基因的cDNA全长1461bp,编码具480个氨基酸残基的前体蛋白,其氨基酸序列与人,牛,大鼠,小鼠同源性达到97%以上.长白猪Akt2基因cDNA全长为1505bp,编码具481个氨基酸残基的前体蛋白,其氨基酸序列与人,牛,大鼠,小鼠的同源性高达97%以上.SMART分析表明,猪Akt1和Akt2蛋白均包含了与PI-3K结合的PH结构域及2个具有丝氨酸/苏氨酸激酶催化活性的S_TKc结构域.RT-PCR检测结果显示:Akt1mRNA在垂体、心脏、肝脏、脾脏、肌肉组织中高表达,在大脑、小脑、肾脏表达丰度较低.Akt2则在小脑、垂体、心脏、肝脏、脾脏和肌肉组织中高表达,而在大脑和肾脏中表达丰度较低.     

不同种类电气石材料对甲醛净化性能及机理研究

利用电子探针和X射线衍射仪, 分析来自新疆(XJ, XJZ, XJF)、云南(YN)、广西(GX)和巴西(BXL)4个产地的6种电气石样品的成分及结构差异, 并研究种类不同、粒度相同的电气石以及种类相同、粒度不同的电气石对甲醛的净化能力。电子探针结果表明, XJ, YN和GX属铁电气石类, 其铁含量依次增加, XJZ为镁电气石, XJF为锂电气石, BXL为钠锰电气石。X射线衍射结果表明, 铁的存在使电气石的晶胞参数变大。实验证实, 当粒度相同时, 不同种类电气石净化甲醛的效果存在较大差异, 属锂电气石的XJF最优, 铁含量最高的GX最次, 6种样品净化甲醛的能力大小为XJF>XJ>XJZ>BXL>YN>GX, 推测与Y八面体的离子占位不同引发的多面体扭曲程度有关; 种类相同的电气石粒度越小, 电学性能越好, 净化甲醛的效率越高。     

一种基于数据划分的家蚕性状比较方法

性状比较的结果数据有助于家蚕优良亲本选择和性状改良工作.人工比较家蚕性状的工作量大,效率低,且易出错.以家蚕茧层率性状比较为例,提出一种基于数据划分的家蚕性状比较方法,把家蚕茧层率数据按比较年季和比较均值分为2类,分别计算性状趋势、性状均值变化率和综合性状均值变化率等成绩,利用线性回归、方差分析等统计方法,比较3个家蚕品种连续10年春蚕期的茧层率.实验结果显示,该方法能够灵活地对任意家蚕品种、任意蚕期数据段比较性状指标,是一种通用的性状比较方法,具有一定的实用性.     

桃蚜可高效率地传播小西葫芦黄化花叶病毒新疆株

研究了桃蚜传播小西葫芦黄化花叶病毒新疆株的规律，结果表明，无毒蚜虫能够获毒的最短取食时间为30s，获毒率随取食时间的延长而提高，取食时间为70s以上时，获毒率达100％；带毒蚜虫传毒的最短取食时间为0.25h，当取食时间超过最短传毒时间时，传毒率随取食时间的增加而升高，取食时间达2h以上时，传毒率达100％；带毒蚜虫能够传毒的最少头数为1头，传毒率随单株植物带毒蚜虫数目的增加而升高，单株虫量达15头以上时，传毒率达100％，说明桃蚜可以高效率地传播小西葫芦黄化花叶病毒新疆株。     

应用梭状芽孢杆菌口服疫苗载体表达HIV p24蛋白的研究

应用一种不产毒的梭状芽孢杆菌（Clostridium perfringens,C.P.）作为口服疫苗载体表达HIVp24蛋白。用PCR扩增HIVp24基因,酶切后插入到含有C.P.序列的pJRC200质粒的CPEORF中,将重组的cp24质粒转化无致病作用的ATCC3624菌株。使HIVp24基因在C.P.形成芽孢时大量表达。结果应用SDS_PAGE和West Blot分析HIVp24在重组C.P.繁殖体和芽孢中的表达情况,发现HIVp24蛋白在C.P.芽孢体中大量表达,而在C.P.繁殖体中却没有表达。证明在体外构建了含有HIVp24基因的重组pJRC200质粒,成功地表达了HIVp24蛋白并得到了HIVp24的高表达ATCC3624菌株。为成功研究C.P.作为一种口服HIV疫苗载体奠定了基础。     

新集一矿孔隙结构特征研究

为进一步了解新集一矿煤层孔隙结构特征,通过压汞法和低温氮等温吸附法实验手段对新集一矿主采煤层孔隙发育特征进行研究。发现新集一矿主采煤层孔隙度较比两淮矿区其他地区高。其中以8煤层8XJ5号样为代表孔隙分布以小孔最发育,大孔较发育,中孔少见为特征和以13-1煤层13XJ10号样为代表孔隙分布以小孔最发育为特征的基质孔,孔隙之间的连通差,不利于瓦斯的抽排;以11-2煤层11XJ3号样和6-1煤层6XJ15号样为代表中孔发育,孔隙以裂隙性为主,孔隙之间的连通较好,有利于瓦斯的运移。     

水霉拮抗菌拮抗活性物质的分离鉴定

本实验采用琼脂扩散法证实,功能拮抗菌株TCCC11322克隆及培养上清,对寄生水霉XJ001生长都有明显的抑制效果,在此实验基础上,对无菌培养上清,进行粗蛋白、粗多糖和肽类物质的分离提取,并对各组分进行了水霉生长拮抗活性检验鉴定,结果证明,培养上清中的分泌性蛋白成分对水霉生长具有明显的抑制效果。     

Improved immunogenicity of a peptide epitope after fusion to hepatitis B core protein

Synthetic vaccines for viral diseases can use defined regions of viral proteins as immunogens: the peptide sequence of amino acids 141-160 of the VP1 protein of foot and mouth disease virus (FMDV) elicits virus-neutralizing antibodies to protect guinea pigs, cattle and pigs either when coupled to a carrier protein or when administered in liposomes or in incomplete Freund's adjuvant. The immune response to these peptides is much lower than that to complete virus particles and the same sequence fused to the N terminus of beta-galactosidase did not produce a more potent immunogen than synthetic peptide alone. We report here an expression system for immunogenic epitopes linked to a carrier protein, hepatitis B core antigen, to form part of a virus-like complex which can present these epitopes to the immune system at high density. The immunogenicity of these structures approaches that of FMDV particles.