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1.
利用反转录-PCR方法,从分泌肾综合症出血热病毒(HFRSV)内影像型抗独特型人单抗杂交瘤细胞系(C8)中,成功地克隆了抗HFRSV1d人单抗重链可变区基因,并将此基因重组入M13噬菌体DNA中,测定了此重链可变区基因全序列。经计算机分析,基因全长共351bp、编码117个氨基酸,氨基酸序列同源性分析发现所推得的该单抗CDR2区与HFRSVG2蛋白C端有同源区,此区可能具有较强的抗原性。  相似文献   

2.
以来自哈尔滨传染性法氏囊病病毒(IBDV) 强毒株(Harbin 毒株,H) 的基因组RNA为模板,用反转录聚合酶链反应(RT- PCR) 的方法得到了其A 节段的全长cDNA 片段,分5'端(1 659bp) 和3'端(1 444bp) 上下两段分别克隆到pGEMB○R - T 载体上,测定了其核苷酸顺序,在长为3 101 bp 中含有两个阅读框ORFA1 和ORFA2 ,分别编码1 012 个氨基酸的前体蛋白(VP2 - 4 -3) 和145 个氨基酸的VP5,ORFA1 和ORFA2 有部分的重叠。将核苷酸序列及推测出的氨基酸序列与已报道的IBDV 血清Ⅰ型和Ⅱ型毒株的相应序列进行了比较,结果表明:H 毒株与其它血清Ⅰ型毒株之间,在核苷酸水平上存在25bp - 267bp 的差异;在氨基酸水平上存在17 ~40 个氨基酸的差异。在VP2 - 4 - 3 内比较显示,H 毒株与P2 、Cu- 1 之间氨基酸的差异最小为1 .7% ,H 毒株与UK661 之间氨基酸的差异最大为3 .9 % 。变异主要发生在VP2 的可变区(206 - 350 位氨基酸) ,在H 毒株所特有的12 个氨基酸当中,该区就占5 个,代表1 .76 % 的变异。VP4、VP3 和VP5区各有  相似文献   

3.
本文测定了粘虫核多角体病毒的一个长3336bp的大片段,该片段包括了3个完整的开放阅读框(ORF2、ORF3、ORF4)和两个不完整的开放阅读框(ORF1、ORF5)。与AcNPV比较发现,ORF1编码重蛋白与P94同源,ORF3、ORF4、ORF5分别与AcNPV的ORF60、ORF59、ORF57同源。且转录方向一致。ORF2则是LsNPV的特有基因。分析结果表明,LsNPV基因组上基因的组织  相似文献   

4.
胡子信  张曼夫 《病毒学报》1999,15(4):330-338
以来自哈尔滨传染性法氏囊病病毒(IBDV)强素株(Harbin 毒株,H)的基因组RNA为模板,用反转录聚合酶链反应(RP-PCR)的方法得到了其A节段的全长cDNA片段,分5端(1659bp)和3端(1444bp)上下两段分别克隆到pGEMB-T载体上,测定了其核苷酸顺序,在长为3101bp中含有两个阅读枢ORF A1和ORF A2,分别编码1012个氨酸酸的前体蛋白(VP2-4-3)和145个  相似文献   

5.
牛泡沫病毒反式激活因子在内部启动子上应答元件的研究   总被引:4,自引:0,他引:4  
泡沫病毒的基因转录依赖至少两个不同的启动子:LTR调节病毒结构蛋白的表达,而内部启动子(IP)则起始调节蛋白mRNA的转录。牛泡沫病毒(BFV)在env与3’LTR之间有两个重叠的开放阅读框架orf-1和orf-2,分别编码BFV ORF-1、ORF-2等多种调节蛋白。这此蛋白中BFV ORF-1为转录激活因子,称为Taso Tas对LTR及IP均有反式激活作用。BFV中第二类启动子IP的存在反映  相似文献   

6.
王勇  王登顺 《遗传学报》1996,23(2):91-95
根据鼠免疫球蛋白重。轻链可变区基因FR1和FR4的序列保守性,化学合成了适于体外扩增Ig重、轻链可变区基因(V_H和V_L)的数对引物。以分泌抗人肺腺癌单抗的杂交瘤细胞株WLA-2C4的基因组DNA为模板,PCR扩增V_H和V_L基因,分别克隆人pUC19载体。转化子经蓝、白斑筛选,酶切鉴定,双脱氧测序证实确为鼠单抗可变区基因,其中V_H基因全长为348bp,编码116aa,属重链ⅡB亚类;V_L基因全长318bp,编码106aa,属K轻链Ⅵ亚类。  相似文献   

7.
新近发现的PRRSV是单股RNA病毒,属于不久前成立的动脉炎病毒科。其基因组包括8个开放阅读框架(ORF),其中ORF5编码糖基化的囊膜(E)蛋白,是病毒的主要免疫保护性抗原之一。为了研制PRRS基因疫苗,扩增并克隆了PRRSV-1a株ORF5,并将其亚克隆至真核表达载体pCR3-Uni的CMV启动子下游,构建成真核表达质粒,为PRRS基因免疫奠定基础  相似文献   

8.
分别利用5’RACE和3’RACE确定了CMVSDRNA2的5’和3’末端序列,在此基础上,利用RTPCR得到了RNA2的5’端一半的cDNA克隆pC25和3’端一半的cDNA克隆pC23,并通过拼接构建了RNA2全长cDNA克隆pC2F。通过对pC25和pC23进行序列测定,得到了RNA2的全序列。序列分析结果表明CMVSDRNA2由3048nt组成,其中存在2个部分重叠的阅读框ORF1(79~2652nt)和ORF2(2414~2746nt),分别编码858aa的2a蛋白和111aa的2b蛋白,并在2a蛋白的序列中发现了动植物病毒复制酶所特有的两个保守序列。该株系RNA2核苷酸序列与分属CMVI亚组的Fny株系和II亚组的Q株系RNA2的核苷酸序列同源性分别为917%和756%;2a蛋白的氨基酸序列同源性分别为938%和677%,2b蛋白的氨基酸序列同源性分别为830%和513%。同源性比较的结果表明SD株系属于CMVI亚组。  相似文献   

9.
减蛋综合征病毒100K蛋白基因的克隆与序列分析   总被引:2,自引:0,他引:2  
用常规方法提取减蛋综合征病毒(EDSV)中国分离株(AA2株)病毒DNA,分别构建了限制性内切酶HindⅢ、SphⅠ、PstⅠ水解片段的全基因文库,并对其中100K蛋白基因的序列进行了分析。EDSV100K蛋白基因位于减蛋综合征病毒基因组55.7~64.8物理图谱单位(m.u),共2091个核苷酸(nt),其编码产物由696个氨基酸(aa)组成,推测其分子量为77.7kD。编码蛋白氨基酸同源性分析表明,EDSV100K蛋白与人腺病毒(Ad2、Ad5、Ad12、Ad41)、Ⅰ群禽腺病毒(CELO和FAV10)的同源性为32.3~34.4%之间,而与羊腺病毒(OAV)的同源性达到56.4%。  相似文献   

10.
应用基因工程的方法,将含有巨细胞病毒(CMV)启动子的基因片段和人粒细胞-巨噬细胞集落刺激因子(hGM-CSF)的cDNA,克隆进逆转录病毒载体N2A,得到重组质粒N2A/CMV/hGM-CSF.经脂质体包装并转染包装细胞,通过G418药物筛选,得到抗性克隆。经PCR和Southemblot检测证实,GM-CSF基因已整合到该克隆细胞的染色体上,获得的逆转录病毒滴度达10 ̄4CFU/ml,克隆细胞培养上清用TF-1细胞可检测到GM-CSF活性。  相似文献   

11.
R. Singh    S. K. Raj    V. Prasad 《Journal of Phytopathology》2008,156(4):222-228
A Begomovirus causing yellow vein mosaic disease of pumpkin (Cucurbita maxima L.) was characterized at molecular level by cloning and sequence analysis of its complete DNA‐A genome. The DNA‐A of the isolate contains 2758 nucleotides which encode six open reading frames (ORFs): AV1 and AV2 in the virion‐sense and AC1, AC2, AC3 and AC4 in the complementary‐sense. Based on the highest (96%) sequence identities and close phylogenetic relationships with Squash leaf curl China virus species, the Begomovirus was identified as strain of Squash leaf curl China virus. The presence of DNA‐B genome of the virus strain was also detected by dot blot hybridization test using DNA‐B specific probe.  相似文献   

12.
Reports on the genetic variability and evolution of natural populations of DNA viruses are scarce in comparison with the abundant information on the variability of RNA viruses. Geminiviruses are plant viruses with circular ssDNA genomes that are replicated by the host plant DNA polymerases. Whitefly-transmitted geminiviruses (WTG) are the agents of important diseases of crop plants and best exemplify emerging plant viruses. In this report we have analyzed the genetic diversity of cotton leaf curl geminivirus (CLCuV), a typical emerging WTG. No genetic differentiation was observed between isolates from different host plant species or geographic regions. Thus, the analyzed isolates represented a unique, undifferentiated population. Genetic variability, estimated as nucleotide diversities at synonymous positions in open reading frames (ORFs) for the AC1 (=replication) protein and coat protein (CP = AV1), was very high, exceeding the values reported for different genes in several plant and animal RNA viruses. This was unexpected in a virus that uses the DNA replication machinery of its eukaryotic host. Diversities at nonsynonymous positions, on the other hand, indicated that variability may be constrained in the genome of CLCuV. The ratio of nonsynonymous-to-synonymous substitutions varied for the different ORFs: they were higher for CP than for AC1 and lower still for the AC4 and AV2 ORFs, which overlap AC1 and CP ORFs, respectively. Analysis of nucleotide diversities at synonymous and nonsynonymous positions of the AC4 and AV2 ORFs suggest that their evolution is constrained by AC1 and CP, respectively. Data suggest that AC4 and AV2 are new genes that may have originated by overprinting on the preexistent AC1 and CP genes. Evidence for recombination was found for the AC1 and CP ORFs and for the noncoding intergenic region (IR). Data indicate that the origin of replication is a major recombination point in the IR, but not the only one. Analyses of the IR also suggest that recombinants may be frequent in the population and that recombination may have an important role in the generation of CLCuV variability. Received: 26 February 1999 / Accepted: 31 May 1999  相似文献   

13.
《Gene》1997,185(2):181-186
Bovine adenovirus type 2 (BAV2) is a medium size double-stranded DNA virus which infects both bovine and ovine species, resulting in mild respiratory and gastrointestinal disorders. To better understand the virus and its growth characterisitics in Madin-Darby bovine kidney (MDBK) cells, we have cloned and sequenced the extreme right-end segment of the BAV2 genome (90.5–100 map units). Analysis of the nucleotide sequence revealed 40 potential open reading frames (ORFs) with coding capacity for polypeptides that are 25 or more amino acid (aa) residues long. Six of these ORFs encode polypeptides that show homology to well-characterized early region 4 (E4) proteins of human adenovirus type 2 (Ad2) and Ad12. ORF1 has the potential to encode a 114 aa long polypeptide that is 54% homologous to the E4 14 kDa protein of Ad2. ORF2 encodes a 78 aa long polypeptide that exhibits 40% homology to the E4 13 kDa protein of Ad2. ORFs 3–6 encode polypeptides that have homology to the E4 34 kDa protein encoded by ORF6 of Ad2 and Ad12. ORFs 3, 4 and 5 encode 128, 96 and 31 aa long polypeptides, respectively. The 128-aa polypeptide exhibits 59% homology, while the 96 and 31 aa long polypeptides exhibit 61% and 70% homology to the E4 34 kDa protein, respectively. ORF6 has the potential to encode a 57 aa long polypeptide that has 67% homology to the E4 34 kDa protein of Ad2 and 50% homology to the E4 34 kDa protein of Ad12.  相似文献   

14.
广东番茄曲叶病毒G2分离物基因组DNA-A的分子特征   总被引:4,自引:0,他引:4  
从采集于广东的番茄曲叶病病株上分离到病毒分离物G2 ,序列分析结果表明 ,其DNA_A为单链环状 ,全长2 74 4nt,共有 6个ORF ,其中病毒链上编码AV1(CP)、AV2 ,互补链上编码AC1、AC2、AC3和AC4。BLAST结果显示 ,与G2基因组有同源关系的病毒均属双生病毒科菜豆金色花叶病毒属。序列比较结果显示 ,G2与菜豆金色花叶病毒属病毒的DNA_A序列同源率均不超过 83% ,其中同源率最高的是PaLCuCNV_[G10 ](82 8% )。进一步比较发现 ,它们的基因间隔区 (IR)变异最大 (同源率为 30 9%~ 81 8% ) ;CP氨基酸序列的同源率较高 (77 6 %~ 99 2 % ) ,AC4蛋白氨基酸序列的同源率较低 (4 3 5 %~ 78 8% )。系统进化关系分析结果也显示 ,G2与已报道的菜豆金色花叶病毒属病毒的亲缘关系均较远。因此 ,G2可能是双生病毒科菜豆金色花叶病毒属中一个未报道的新种 ,命名为广东番茄曲叶病毒 (TomatoleafcurlGuangdongVirus ,ToLCGDV)  相似文献   

15.
To localize gene that may encode immunogens potentially important for recombinant vaccine design, we have analysed a region of the equine herpesvirus type-1 (EHV-1) genome where a glycoprotein-encoding gene had previously been mapped. The 4707-bp BamHI-EcoRI fragment from the short unique region of the EHV-1 genome was sequenced. This sequence contains three entire open reading frames (ORFs), and portions of two more. ORF1 codes for 161 amino acids (aa), and represents the C terminus of a possible membrane-bound protein. ORF2 (424 aa) and ORF3 (550 aa) are potential glycoprotein-encoding genes; the predicted aa sequences contain possible signal sequences, N-linked glycosylation sites and transmembrane domains; they also show homology to the glycoproteins gI and gE of herpes simplex virus type-1 (HSV-1), and the related proteins of pseudorabies virus and varicella-zoster virus. The predicted aa sequence of ORF4 shares no homology with other known herpesvirus proteins, but the nucleotide sequence shows a high level of homology with the corresponding region of the EHV-4 genome. ORF5 may be related to US9 of HSV-1.  相似文献   

16.
A significant number of mycoviruses have been identified that are related to plant viruses, but their evolutionary relationships are largely unexplored. A fusarivirus, Rhizoctonia solani fusarivirus 4 (RsFV4), was identified in phytopathogenic fungus Rhizoctonia solani (R. solani) strain XY74 co-infected by an alphaendornavirus. RsFV4 had a genome of 10,833 nt (excluding the poly-A tail), and consisted of four non-overlapping open reading frames (ORFs). ORF1 encodes an 825 aa protein containing a conserved helicase domain (Hel1). ORF3 encodes 1550 aa protein with two conserved domains, namely an RNA-dependent RNA polymerase (RdRp) and another helicase (Hel2). The ORF2 and ORF4 likely encode two hypothetical proteins (520 and 542 aa) with unknown functions. The phylogenetic analysis based on Hel2 and RdRp suggest that RsFV4 was positioned within the fusarivirus group, but formed an independent branch with three previously reported fusariviruses of R. solani. Notably, the Hel1 and its relatives were phylogenetically closer to helicases of potyviruses and hypoviruses than fusariviruses, suggesting fusarivirus Hel1 formed an evolutionary link between these three virus groups. This finding provides evidence of the occurrence of a horizontal gene transfer or recombination event between mycoviruses and plant viruses or between mycoviruses. Our findings are likely to enhance the understanding of virus evolution and diversity.  相似文献   

17.
IS421, a new insertion sequence in Escherichia coli   总被引:2,自引:0,他引:2  
The nucleotide sequence of a new insertion sequence (IS) in Escherichia coli, IS421, was determined. It is 1340 bp long and contains inverted repeats of 22 bp at its termini. It is flanked by 13 bp direct repeats apparently generated upon insertion. There are two ORFs longer than 200 bp in IS421. One can encode a polypeptide of 371 amino acids (aa) and the other, which is on the other strand, can encode a polypeptide of 102 aa. The C-terminal part of the 371 aa polypeptide shows some homology to that of transposases encoded in some other known IS elements. The copy number of IS421 in chromosomal DNA was 4 for E. coli K-12 and B, and 5 for E. coli C, as determined by the Southern hybridization of restriction fragments.  相似文献   

18.
19.
Mulberry vein banding associated virus (MVBaV) that infects mulberry plants with typical vein banding symptoms had been identified as a tentative species of the genus Tospovirus based on the homology of N gene sequence to those of tospoviruses. In this study, the complete sequence of the tripartite RNA genome of MVBaV was determined and analyzed. The L RNA has 8905 nucleotides (nt) and encodes the putative RNA-dependent RNA polymerase (RdRp) of 2877 aa amino acids (aa) in the viral complementary (vc) strand. The RdRp of MVBaV shares the highest aa sequence identity (85.9%) with that of Watermelon silver mottle virus (WSMoV), and contains conserved motifs shared with those of the species of the genus Tospovirus. The M RNA contains 4731 nt and codes in ambisense arrangement for the NSm protein of 309 aa in the sense strand and the Gn/Gc glycoprotein precursor (GP) of 1,124 aa in the vc strand. The NSm and GP of MVBaV share the highest aa sequence identities with those of Capsicum chlorosis virus (CaCV) and Groundnut bud necrosis virus (GBNV) (83.2% and 84.3%, respectively). The S RNA is 3294 nt in length and contains two open reading frames (ORFs) in an ambisense coding strategy, encoding a 439-aa non-structural protein (NSs) and the 277-aa nucleocapsid protein (N), respectively. The NSs and N also share the highest aa sequence identity (71.1% and 74.4%, respectively) with those of CaCV. Phylogenetic analysis of the RdRp, NSm, GP, NSs, and N proteins showed that MVBaV is most closely related to CaCV and GBNV and that these proteins cluster with those of the WSMoV serogroup, and that MVBaV seems to be a species bridging the two subgroups within the WSMoV serogroup of tospoviruses in evolutionary aspect, suggesting that MVBaV represents a distinct tospovirus. Analysis of S RNA sequence uncovered the highly conserved 5’-/3’-ends and the coding regions, and the variable region of IGR with divergent patterns among MVBaV isolates.  相似文献   

20.
Molecular Evolution of the Genomic RNA of Apple Stem Grooving Capillovirus   总被引:1,自引:0,他引:1  
The complete genome of the German isolate AC of Apple stem grooving virus (ASGV) was sequenced. It encodes two overlapping open reading frames (ORFs), similarly to previously described ASGV isolates. Two regions of high variability were detected between the ASGV isolates, variable region 1 (V1, from amino acids (aa) 532 to 570), and variable region 2 (V2, from aa 1,583 to 1,868). The phylogenetic analysis of the V1 and V2 regions suggested that the ASGV diversity was structured by host plant species rather than geographical origin. The dN/dS ratio between nonsynonymous and synonymous nucleotide substitution rates varied greatly along the ASGV genome. Most of ORF1 showed predominant negative selection except for the two regions V1 and V2. V1 showed an elevated dN and an average dS when compared to the ORF1 background but no significant positive selection was detected. The V2 region of ORF1 showed an elevated dN and a low dS when compared to the ORF1 background with an average dN/dS????3.0 indicative of positive selection. However, the V2 area includes overlapping ORFs, making the dN/dS estimate biased. Joint estimates of the selection intensity in the different ORFs by a recent method indicated that this region of ORF1 was in fact evolving close to neutrality. This was convergent with previous results showing that introduction of stop codons in this region of ORF1 did not impair plant infection. These data suggest that the elimination of a stop codon caused the overprinting of a novel coding region over the ancestral ORF.  相似文献   

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