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猪瘟病毒NS4B蛋白是猪瘟病毒编码的一个非结构蛋白,由347个氨基酸残基组成。NS4B在病毒中可能与致细胞病变有关,具体功能尚不清楚。本试验利用PCR扩增NS4B基因,利用双酶切、连接和转化等操作方法,与质粒载体p EGFP-N3构建成真核表达重组质粒,双酶切检测猪瘟病毒NS4B真核表达载体构建成功。测定重组质粒序列,并验证表达成功,为今后的功能研究奠定了基础。 相似文献
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禽流感病毒非结构蛋白的研究进展 总被引：1，自引：0，他引：1
流感病毒含8个负链单股独立的RNA片段,共编码10种蛋白。每种蛋白具有不同的结构和功能。其中NS基因合成NS1和NS2两种蛋白质,NS1为非结构蛋白,NS2为结构蛋白,这两种蛋白质大量存在于感染的细胞中。NS1蛋白在病毒转录及感染过程中起重要作用,与禽流感病毒的致病性密切相关,其主要功能是以多种方式解除宿主干扰素防御系统。随着分子生物技术的快速发展,关于NS1蛋白在禽流感病毒致病性方面的作用的研究也取得了一定的进展。文章主要对流感病毒NS基因的特点及其编码蛋白质功能进行综述。 相似文献
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《畜牧与兽医》2016,(7):132-135
NS1蛋白不存在于病毒粒子中,但在感染A型流感病毒的细胞中表达量很高,因此,人们普遍认为NS1蛋白是A型流感病毒的非结构蛋白。NS1蛋白是一种相对较小的多肽,在抗病毒作用中能够选择性地增强病毒mRNA的翻译,并且可以调节病毒RNA的合成;在流感病毒感染的先天性免疫应答中,NS1是主要的病毒拮抗剂,通过在IFN系统中阻止关键因子的激活发挥作用;在调整PI3K信号通路中NS1通过显著性的影响总体的基因表达来破坏宿主细胞的内环境稳定,从而影响细胞凋亡。因此,NS1蛋白在克服机体阻挡病毒感染的第一道防线中的作用至关重要,是影响流感病毒致病机理和适应宿主的一个重要毒力因子。本文对NS1蛋白的结构及功能进行了概述。 相似文献
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日本乙誓脑炎病毒NS1基因及其疫苗的研究进展 总被引：1，自引：0，他引：1
日本乙型脑炎是由日本乙型脑炎病毒引起的严重的人畜共患传染病。乙型脑炎病毒的三种糖基化蛋白PrM、E和NS1是其主要免疫保护性抗原,其中PrM和E蛋白是日本乙型脑炎病毒的结构蛋白,其结构和相关疫苗研究的报道较多。NS1蛋白是日本乙型脑炎病毒的非结构蛋白,其主要参与病毒复制的早期阶段,推测可能参与病毒组装和释放。可诱导补体依赖性溶细胞反应,不能产生中和抗体,能诱发机体在非中和性抗体存在的情况下产生保护性免疫。在接种乙脑病毒的细胞的细胞内、细胞表面及上清液中均含有大量的NS1蛋白。对小鼠接种NS1蛋白或NS1蛋白特异性抗体,均能让小鼠产生对乙脑病毒感染的保护性免疫作用。本文主要就乙型脑炎病毒非结构基因NS1及其免疫疫苗的研究进展进行综述,为其更进一步深入研究IEV NS1基因及其相关疫苗奠定基础。 相似文献
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《中国兽医杂志》2020,(4)
为表达蓝舌病病毒NS3蛋白,将蓝舌病病毒血清8型NS3序列克隆至pFastBac-HTA载体上,经转座、转染后获得重组杆状病毒rBac-NS3;用间接免疫荧光(IFA)检测NS3蛋白的表达情况,并采用亲和层析进行纯化;以Western Blot和间接ELISA方法鉴定NS3蛋白的反应原性。IFA结果显示,重组杆状病毒rBac-NS3感染的Sf9细胞可以大量表达NS3蛋白。Western Blot结果显示,纯化后的NS3蛋白与抗His单抗和绵羊蓝舌病阳性血清能够发生特异性反应;以纯化的NS3蛋白为包被抗原建立的间接ELISA方法可以区分蓝舌病阳性血清和阴性血清,进一步验证本试验利用杆状病毒表达系统成功表达并纯化出蓝舌病病毒NS3蛋白,表明纯化后的NS3蛋白具有良好的反应原性,可作为开发鉴别诊断试剂盒的备选蛋白及NS3蛋白的结构和生物学功能研究。 相似文献
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The bovine viral diarrhea virus (BVDV) strains exist as two biotypes, cytopathic (cp) and noncytopathic (ncp), according to their effects on tissue culture cells. It has been previously reported that cell death associated to cp BVDV in vitro is mediated by apoptosis. Here, experiments were conducted to determine the involvement of the NS3 protein in the induction of apoptosis. The NS3- and NS3Delta50 (deleted from the NH2-terminal 50 amino acids)-cDNA encoding sequences of BVDV NADL cp reference strain were cloned into adenoviral vectors (AdV) from which the BVDV gene of interest could be expressed from a tetracycline-responsive promoter. A549tTA cells infected in vitro with NS3 or NS3Delta50-expressing AdV showed cytopathic changes characterized by cell rounding and detachment, and nucleus chromatin condensation. DNA fragmentation assays, cytochrome c release, and activation of cellular caspases performed on these infected cells clearly correlated with the observed cytopathic changes with apoptosis. The BVDV NS3Delta50-induced apoptotic process was inhibited by caspase-8- and -9-specific peptide inhibitors (Z-IETD-FMK and Z-LEHD-FMK). Furthermore, apoptosis was inhibited in cells expressing the R1 subunit of herpes simplex virus type 2 ribonucleotide reductase (HSV2-R1) or hsp70, two proteins which are known to inhibit apoptosis associated with caspase-8 activation and cytochrome c release-dependent caspase-9 activation, respectively. Given that HSV2-R1, a specific inhibitor of the caspase-8 activation pathway, efficiently suppressed apoptosis and also prevented caspase-9 activation, the overall results indicate that the BVDV NS3/NS3Delta50 induces apoptosis initiated by caspase-8 activation and subsequent cytochrome c release-dependent caspase-9 activation. 相似文献
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牛病毒性腹泻病毒研究进展 总被引：2，自引：0，他引：2
牛病毒性腹泻病毒是黄病毒科瘟病毒属的代表种,主要感染牛并引发疾病,造成严重的经济损失.自该病毒发现至今,已有60余年研究历史,随着生物学理论及技术不断发展,特别是反向遗传技术以及蛋白组学在病毒研究领域的应用,人们对该病毒产生了一些新的认识.论文从牛病毒性腹泻病毒的病毒粒子结构组成及功能、病毒复制周期及蛋白裂解过程、细胞损伤及免疫逃逸机制三个方面阐述了近几年牛病毒性腹泻病毒分子生物学研究的进展,重点讨论了病毒复制周期、多聚蛋白裂解、致细胞病变效应和免疫逃逸机制. 相似文献
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近年来大量的研究结果表明,甲型流感病毒的非结构蛋白质1(NS1)是在感染病毒期间具有多个辅助功能的毒力因子,为揭示该人畜共患病毒的致病性机理提供重要参考价值。这篇文章综述了该病毒蛋白质一些最新研究进展,包括NS1的合成、结构、亚细胞定位及在病毒复制机制、宿主天然免疫应答或/和适应性免疫应答、细胞信号传导中发挥各种作用。作者重点阐述了NS1-RNA之间和NS1-蛋白质之间的相互作用,因为它们都是上述功能和过程的基础。同时突出不同毒株NS1蛋白可能特有的特性,鉴于此,也讨论了NS1对人和动物甲型流感病毒致病性的某些作用。最后概述NS1可能应用于流感疫苗的研制、新抗病毒药物的研发等。同时比较了乙、丙型流感病毒NS1研究新进展。 相似文献
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Suppression of bovine viral diarrhea virus replication by small interfering RNA and short hairpin RNA-mediated RNA interference 总被引：2，自引：0，他引：2
Bovine viral diarrhea virus (BVDV) is a ubiquitous viral pathogen that affects cattle herds' worldwide causing significant economic loss. The current strategies to control BVDV infection include vaccination (modified-live or killed) and control of virus spread by enhanced biosecurity management, however, the disease remains prevalent. With the discovery of the sequence-specific method of gene silencing known as RNA interference (RNAi), a new era in antiviral therapies has begun. Here we report the efficient inhibition of BVDV replication by small interfering (siRNA) and short hairpin RNA (shRNA)-mediated gene silencing. siRNAs were generated to target the 5' non-translated (NTR) region and the regions encoding the C, NS4B and NS5A proteins of the BVDV genome. The siRNAs were first validated using an EGFP/BVDV reporter system and were then shown to suppress BVDV-induced cytopathic effects and viral titers in cell culture with surprisingly different activities compared to the reporter system. Efficient viral suppression was then achieved by bovine 7SK-expressed BVDV-specific shRNAs. Overall, our results demonstrated the use of siRNA and shRNA-mediated gene silencing to achieve efficient inhibition of the replication of this virus in cell culture. 相似文献
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Khalid Munir 《Journal of veterinary science (Suw?n-si, Korea)》2006,7(2):167-176
We have previously described the development of a one-tube SYBR Green real-time RT-PCR assay for the detection and quantitation of infectious salmon anemia virus (ISAV) in various biological samples. The twofold aim of the present study was to verify that the optimized SYBR Green real-time RT-PCR conditions could detect ISAV isolates of different geographic origins, and to analyze the growth patterns of the selected ISAV isolates in the Chinook head salmon embryo (CHSE)-214 cells by this assay to better characterize their CHSE-phenotypes. A total of 24 ISAV isolates were used in this study. The results indicated that the SYBR Green real-time RT-PCR could detect ISAV of different geographic origins or laboratory sources. The capacity of ISAV isolates to cause cytopathic effects (CPE) in the CHSE-214 cell line, viral titration of the infected CHSE-cell harvests, and analysis of viral RNA levels in CHSE-214 cells at post-infection day zero, 7 and 14 by SYBR Green real-time RT-PCR confirmed the existence of three CHSE-phenotypes of ISAV: replicating cytopathic, replicating non-cytopathic, and non-replicating non-cytopathic. The identification of these three CHSE-phenotypes of ISAV has important implications from diagnostic and biological points of view. 相似文献
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为分离鉴定新疆地区牛病毒性腹泻-黏膜病病毒(BVDV)流行毒株,掌握该毒株的生物学特性,本试验在新疆北疆部分地区采集牛病毒性腹泻-黏膜病(BVD/MD)疑似病例的粪便,通过RT-PCR检测、细胞分离培养、间接免疫荧光抗体检测、免疫电镜观察及血清中和试验5种方法对毒株进行分离和鉴定。对毒株的TCID50测定后,再对毒株分别进行乙醚敏感性试验、氯仿敏感性试验、胰蛋白酶敏感性试验、酸碱度敏感性试验、温度敏感性试验及核酸分型试验等理化特性检测。经RT-PCR诊断,病料在286 bp处出现了目的片段。将RT-PCR诊断为阳性的粪便,接种于密度约为80%的单层MDBK细胞出现了细胞病变,盲传5代至出现典型的细胞病变。将F5代细胞培养物采用间接免疫荧光抗体检测,结果产生了与C24V标准毒株相同的特异性黄绿色荧光。免疫电镜观察到了大量呈球形的BVDV粒子,大小20~40 nm。血清中和试验中抗体阳性血清处理组细胞均未出现细胞病变,病毒完全被抗体阳性血清中和。综合以上方法确定分离株为BVDV毒株。对分离株进行毒价和理化特性测定,该毒株TCID50为10-4.5/0.1 mL,对乙醚和氯仿敏感,对胰蛋白酶敏感,耐碱不耐酸,对温度敏感,经54 ℃ 1 h完全被灭活,属于RNA病毒。本试验成功分离到一株新疆BVDV流行毒株,掌握了该毒株的生物学特性,为今后该病的诊断和防控奠定了基础。 相似文献
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D Deregt K G Loewen 《The Canadian veterinary journal. La revue veterinaire canadienne》1995,36(6):371-378
Bovine viral diarrhea virus continues to produce significant economic losses for the cattle industry and challenges investigators with the complexity of diseases it produces and the mechanisms by which it causes disease. This paper updates and attempts to clarify information regarding the roles of noncytopathic and cytopathic bovine viral diarrhea viruses in persistent infections and mucosal disease. It also covers, in brief, what is known of the new diseases: thrombocytopenia and hemorrhagic disease, and a disease resembling mucosal disease that is apparently caused solely by noncytopathic virus. Although a good understanding of the roles of the 2 biotypes in the production of persistent infections and the precipitation of mucosal disease has been obtained, there are still unanswered questions regarding the origin of cytopathic viruses and the mechanism by which they cause pathological changes in cells. It is apparent, however, that cytopathic bovine viral diarrhea viruses arise by mutation of noncytopathic viruses, and it is known that p80 is the marker protein for cytopathic viruses. The previous distinction between mild bovine viral diarrhea and fatal mucosal disease has been eroded with the emergence of new virulent bovine viral diarrhea viruses. The new diseases pose a threat to the cattle industry and present a new challenge for investigators. Index Veterinarius (1984-1994) and Medline (1985-1994) databases and personal files updated since 1987 from BIOSIS Previews and Biosciences Information Services were used to search the literature. 相似文献
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Cellular insertions in the NS2-3 genome region of cytopathic bovine viral diarrhoea virus (BVDV) isolates 总被引：1，自引：0，他引：1
When compared to noncytopathic (ncp) bovine viral diarrhoea virus (BVDV), some cytopathic (cp) BVDV contain additional sequences in the NS2-3 genomic region. One of these insertions, which is 270 nucleotides long and of host origin (cINS), was first described for strain NADL. To find out how frequently this type of insertion occurs in other cp BVDV, 32 cp BVDV field isolates and the BVDV reference cp strain Indiana were screened using RT-PCR which detected cINS in NADL. For most cp viruses an RT-PCR product of 402bp indicated the presence of NS2-3 genes without insertions. In addition, one or two DNA fragments, around 600-850bp in size, were amplified from the genomes of 13 cp viruses indicating the presence of insertions. Sequencing of the PCR products, i.e. 402bp DNA fragment (with no insertion) and longer fragments (with insertion) revealed the location of the insertions in the NS2-3 coding region of eight cp BVDV genomes. All of the insertions were confirmed to be of the cINS type and were located in a very similar position to that found previously in the NADL genome. They were in the same reading frame as the viral polypeptide and they encoded 90-140 amino acids. The 5' and 3' ends of the insertions were different in most of the cp isolates studied. Interestingly, a 14-amino-acid stretch at the 5'-end of the insertion in the cp 5569 isolate as well as 15 amino acids at the 3'-end of the insertion in the cp 5.19516 isolate were not homologous to the cINS sequence. No significant matches for these stretches were found in the EMBL and Swissprot databases. 相似文献