共查询到19条相似文献，搜索用时 125 毫秒
1.
合肥地区发生的西瓜花叶病的病原鉴定 总被引：1，自引：1，他引：0
从合肥市郊区的西瓜病叶上分离出一病毒分离物,该分离物的热钝化温度为60~65℃,稀释终点为10-4~10-5,寄主体外存活期为20~23 d,电镜观察病毒粒子约750 nm×13 nm。参考已发表的小西葫芦黄化花叶病毒(Zucchini yellow mosaic virus,ZYMV) CP基因序列设计引物,RT-PCR扩增得到CP基因片段。将该CP片段克隆到pGEM-3Zf (+)中,测序结果表明,该CP基因全长840 nt,共编码279个氨基酸,与国内报道的ZYMV CP基因的核苷酸序列同源性为83.0%~97.3%,氨基酸序列的同源性为91.8%~99.8%,证明该分离物为ZYMV。 相似文献
2.
在北京东郊自然感病的南瓜Cucurbita moschata上获得一病毒分离物(BJ-1),经生物学、血清学和分子生物学鉴定,确定为小西葫芦黄花叶病毒(Zucchini yellowmosaic virus,ZYMV)。为分析其基因组3′端特性,以发病叶片中提取的总RNA为模板,对基因组3′端进行RT-PCR扩增,产物克隆到pMD18-T载体上进行序列分析,共测定了该病毒分离物包括全部CP基因在内的1269bp。该分离物CP基因由837个核苷酸组成,编码279个氨基酸。对包括该分离物在内的30个序列的760bp(含NIb基因3′端56bp和CP基因中的704bp)片段、NIb蛋白与CP蛋白的切割位点、蚜传必需基序的变异、寄主来源及地域来源进行了分析。结果表明,ZYMV不同分离物的基因分型与上述五个因素无明显关系。 相似文献
3.
甘肃省南瓜及西葫芦小西葫芦黄花叶病毒病鉴定 总被引：1，自引：0，他引：1
利用双抗夹心酶联免疫吸附测定(DAS-ELISA)的方法对甘肃出入境南瓜、西葫芦种子及采自河西地区显症病株叶片进行检测,在种子及病叶组织中均检测到ZYMV病毒,其中南瓜种子带毒批次占12.5%,西葫芦种子带毒批次占11.8%。根据已报道的小西葫芦黄花叶病毒(Zucchini yellow mosaic virus)基因组核苷酸序列,设计引物扩增其外壳蛋白(CP)基因,以ELISA阳性种子或病叶组织总RNA为模板,进行RT-PCR扩增,对预期大小的扩增产物进行测序,结果表明扩增获得的核苷酸序列与世界各地的ZYMV分离物CP基因具有高度一致性,综合ELISA检测和RT-PCR的结果,确定南瓜、西葫芦种子可携带ZYMV,且ZYMV是侵染甘肃瓜类作物的重要病毒种类。 相似文献
4.
小西葫芦黄花叶病毒北京分离物的鉴定及其基因组3''端特性分析 总被引：4，自引：1，他引：4
在北京东郊自然感病的南瓜Cucurbita moschata上获得一病毒分离物（BJ-1），经生物学、血清学和分子生物学鉴定，确定为小西葫芦黄花叶病毒（Zucchini yellow mosaic virus，ZYMV）。为分析其基因组3’端特性，以发病叶片中提取的总RNA为模板，对基因组3’端进行RT-PCR扩增，产物克隆到pMD18-T栽体上进行序列分析，共测定了该病毒分离物包括全部CP基因在内的1269bp。该分离物CP基因由837个核苷酸组成，编码279个氨基酸。对包括该分离物在内的30个序列的760bp（含NIb基因3’端56bp和CP基因中的704bp）片段、NIb蛋白与CP蛋白的切割位点、蚜传必需基序的变异、寄主来源及地域来源进行了分析。结果表明，ZYMV不同分离物的基因分型与上述五个因素无明显关系。 相似文献
5.
甘薯潜隐病毒外壳蛋白基因的克隆、表达及其抗血清的制备 总被引：2，自引：0，他引：2
根据已报道的甘薯潜隐病毒(Sweet potato latent virus,SPLV)外壳蛋白(CP)基因的核苷酸序列合成引物,利用RT-PCR方法克隆了SPLV河南分离物(SPLV-HN)的CP基因及部分3'端非编码区序列,序列分析表明,SPLV-HN CP基因由879个核苷酸组成(GenBank登录号为DQ399862),编码293个氨基酸残基。与GenBank中SPLV-CH(X84011)和SPLV-T(X84012)分离物的核苷酸序列相似性分别为96.8%和93.0%;与日本分离物(E15420)的核苷酸序列相似性为83.6%。将CP基因克隆到原核表达载体pET-30a(+)上,SDS-PAGE分析表明,经IPTG诱导,CP基因在大肠杆菌BL21(DE3)pLysS中得到了高效表达。以表达的蛋白为抗原,免疫家兔,制备了SPLV外壳蛋白的特异性抗血清。ACP-ELISA检测结果表明,制备的抗血清可用于田间甘薯样品的检测。 相似文献
6.
西瓜花叶病毒2号南瓜分离物的鉴定及其外壳蛋白基因序列分析 总被引：5，自引：1，他引：5
利用电镜和酶联免疫吸附测定法(ELISA)在黑龙江省采集的南瓜病样中检测到西瓜花叶病毒2号(WMV-2)。再利用免疫PCR (IC-PCR)和反转录PCR (RT-PCR)方法,扩增获得其外壳蛋白(CP)基因片段,并克隆到pGEM-T载体中。核苷酸序列测定表明,该分离物CP基因全长为852个核苷酸,编码由284个氨基酸组成的31.8 kDa蛋白。与国外已报道的WMV-2 CP基因相比,其核苷酸序列同源性为92.2%~94.0%,由此推导的氨基酸序列同源性为94.5%~98.1%。与国内2个分离物相比,和山西分离物核苷酸和氨基酸的同源性都达到98.5%,和郑州分离物核苷酸和氨基酸的同源性分别为91.5%和95.0%。 相似文献
7.
8.
河南省地黄病毒病初步鉴定 总被引：11，自引：0，他引：11
利用血清学、RT-PCR并结合核苷酸序列测定等方法,对河南省地黄病毒病进行了初步鉴定。结果表明,烟草花叶病毒(TMV)为侵染地黄的主要病毒;对TMV地黄分离物(TMV-RH) CP基因的序列分析结果表明,TMV-RH与TMV-U1株系CP基因的核苷酸同源性为86.5%,氨基酸同源性为94.3%;与已发表的TMV其它株系CP基因的核苷酸同源性在76.3%~88.5%之间,氨基酸同源性在79.3%~95.0%之间,同源性较低。根据不同株系CP的氨基酸序列进化树分析,推测该分离物可能为TMV的一个新株系。 相似文献
9.
甘薯脉花叶病毒外壳蛋白基因在大肠杆菌中的表达及其抗血清的制备 总被引：1，自引：1，他引：0
根据已报道的甘薯脉花叶病毒(Sweet potato vein mosaic virus,SPVMV)外壳蛋白（CP）基因的核苷酸序列合成引物,利用RT-PCR方法克隆了SPVMV河南分离物（SPVMV-HN）基因组3′端1.8 kb的基因片段,包括部分NIb 基因序列和完整的CP基因及3′端非编码区序列(3′UTR)。序列分析表明,SPVMV-HN的CP基因由996个核苷酸组成（GenBank登录号为FJ687211）,编码332个氨基酸残基。与已发表的SPVMV其他分离物相比,其推导的氨基酸序列一致性为95.2%～98.5%,与 SPVMV广东分离物的氨基酸序列一致性为97.9%。将CP基因克隆到原核表达载体pET-28a（+）上,SDS-PAGE分析表明,经IPTG诱导,CP基因在大肠杆菌BL21(DE3) pLysS中得到了高效表达。以表达的蛋白为抗原,免疫家兔,制备了SPVMV外壳蛋白的特异性抗血清。ACP-ELISA检测结果表明,制备的抗血清可用于田间甘薯样品的检测。利用SPVMV的抗血清,对采自全国14个省（市）的田间甘薯样品以及嫁接的巴西牵牛样品进行了检测,结果表明,SPVMV在我国甘薯上普遍存在。 相似文献
10.
从山东省感病的烟草上分离获得了一烟草蚀纹病毒(TEV)分离物,命名为TEV-SD1.根据已报道烟草蚀纹病毒外壳蛋白(coat protein CP)基因序列设计并合成2条引物,通过RT-PCR扩增得到长度约800bp的目的片段.将目的片段与质粒pET-22b( )连接,构建了包含TEV CP基因的原核表达载体pETEV-CP,并导入大肠杆菌BL21中.序列分析表明:TEV-SD1的CP基因全长789bp,编码263个氨基酸;与GenBank中已报道的TEV 5个分离物CP基因相比,核苷酸及推导的氨基酸序列同源性为94.1%～98.6%.37℃培养条件下,BL21/pETEV-CP经IPTG诱导表达,SDS-PAGE结果显示,表达的TEV CP融合蛋白的相对分子质量约为33 kDa.以表达的融合蛋白为抗原,免疫家兔,制备的抗血清的效价为1/2048,且具有良好的特异性. 相似文献
11.
甘蔗花叶病毒福建分离物外壳蛋白基因的克隆及序列分析 总被引：3，自引：0，他引：3
A fujian isolate of Sugarcane mosaic virus named SCMV-FJ was isolated from infected sugarcane. Cloning and sequence analysis of the coat protein gene of this isolate was carried out. A pair of primers was designed and synthesized based on the nucleotide sequences of coat protein genes of sugarcane mosaic viruses reported. The coat protein gene of SCMV-FJ was amplified from the extracted total RNA of the infected sugarcane by using RT-PCR, and cloned into the pMD18-T vector. The sequencing result indicated that the cloned segment included a 1137 bp open reading frame(ORF) and a 228 bp 3' untranslated region, in which the ORF comprised the whole coat protein and part of the nuclear inclusion b. The nucleotide and the deduced amino acid sequences of the coat protein gene were compared with those of the other isolates or strains of SCMV subgroup reported in GenBank. The result showed that it shares 56.8%-97.1% and 55.3%-99.4% homology in nucleotide and the putative amino acid sequences, respectively, with the highest amino acid homology of 99.4% with SCMV-D. Thus it was identified as a SCMV-D. This experiment provided a rapid, sensitive and relatively inexpensive method for RT-PCR detection of SCMV. At the same time, the cloning of SCMV-FJ coat protein gene provided the foundation for plant gene engineering against SCMV. 相似文献
12.
Weiland JJ Van Winkle D Edwards MC Larson RL Shelver WL Freeman TP Liu HY 《Phytopathology》2007,97(10):1245-1254
ABSTRACT The first reported U.S. isolate of Beet black scorch necrovirus (BBSV) was obtained and characterized. Host range of the virus for localized and occasionally systemic infection included the Chenopodiaceae and Tetragonia expansa; Nicotiana benthamiana supported symptomless systemic infection by the virus. The complete nucleotide sequence of the genomic RNA of the virus, designated BBSV-Co, exhibits 93% similarity to the genome of the 'Ningxia' isolate of BBSV from China. Amino acid sequence similarity in predicted genes ranged from 95% in the p4 gene to 97% in the p82 and coat protein genes. A potential additional gene exists within the U.S. isolate of BBSV that is absent from Chinese isolates of BBSV due to nucleotide differences between these isolates within the coat protein gene. Coat protein analysis by isoelectric focusing and by mass spectroscopy indicated the presence of phosphorylated residues. Using primer extension analysis of the 5' end of the genome and site-directed mutants of genomic clones of BBSV-Co from which infectious RNA was produced, the native 5' end of the BBSV-Co genome was determined to be 5'-GAAACCTAACC...3', lacking the two terminal adenosine nucleotides in the published sequences of BBSV from China. 相似文献
13.
Biological, serological and molecular characterization of a cucumber mosaic virus isolate from India
A. virus causing mosaic and leaf deformation of Physalis minima has been identified as an isolate of cucumber mosaic virus (CMV) on the basis of its transmission by aphids in a non-persistent manner, polyhedral particles of 29 nm diameter, molecular weight of coat protein subunits us 24-5 kDa. serological relationship with a CMV isolate and a tripartite single-stranded RNA genome with a subgenomic RNA4- Furthermore. cDNA representing coat protein gene was synthesized and cloned. Complete nucleotide sequences (890 nt) were obtained which showed a coat protein gene open reading frame of 657 residues. THE nucleotide sequences provided the 218 amino ACID sequences of the coat protein. Nucleotide as well as amino acid sequences revealed more than 90% identity with the CMV subgroup I strains. 相似文献
15.
Stuart A. MacFarlane Derek J. F. Brown John F. Bol 《European journal of plant pathology / European Foundation for Plant Pathology》1995,101(5):535-539
The coat protein gene of the nematode non-transmissible, SP5 isolate of pea early-browning tobravius was replaced with that of the highly nematode transmissible, PPK20 isolate of tobacco rattle tobravirus. Plants were infected with the recombinant virus when mechanically inoculated and the virus invaded the plants systemically. However, although the PPK20 isolate of TRV was transmitted by nematodes from these plants, the recombinant virus was not transmitted. Therefore, the virus coat protein is not the exclusive determinant of nematode transmission. 相似文献
16.
侵染葫芦的黄瓜绿斑驳花叶病毒广西分离物分子鉴定 总被引：1，自引：0，他引：1
从广西南宁市郊温室大棚中的葫芦[Lagenaria siceraria(Molina)Stand.]上采集到一个表现脉绿、花叶症状的病毒样品,ELISA检测表明,该样品与黄瓜绿斑驳花叶病毒(Cucumber green mottle mosaic virus,CGMMV)有密切的血清学关系,利用RT-PCR方法从样品中扩增获得约500bp的DNA片段,序列分析表明,该片段是CGMMV的外壳蛋白基因,暂将该病毒分离物定名为GX-BG。外壳蛋白基因核苷酸序列系统进化树分析表明,已报道的CGMMV主要分为3大群体,GX-BG与中国辽宁分离物(CGMMV-LN)分别属于不同的群体。 相似文献
17.
葡萄卷叶伴随病毒-3外壳蛋白的原核表达及其特异性抗血清的制备 总被引：2，自引：0，他引：2
利用RT-PCR技术扩增得到了葡萄卷叶伴随病毒-3(Grapevine leafroll associated virus-3,GLRaV-3)中国分离物的外壳蛋白(coat protein,cp)基因。序列分析结果表明,cp基因的长度为942bp,与已报道的其它GLRaV-3分离物的cp核苷酸相似性为91%～99%,编码的氨基酸相似性为95%～100%。将此基因克隆到原核表达载体pET-28a(+)上,转化大肠杆菌BL21(DE3)plysS后用终浓度为1mmol/L的IPTG进行诱导表达,SDS-PAGE及Western blotting分析表明,cp在大肠杆菌中可表达出分子量约为35kDa的蛋白。纯化表达产物后免疫家兔制备抗血清。A蛋白酶联免疫吸附测定(PAS-ELISA)及斑点免疫结合测定(DBIA)结果显示,制备的特异抗血清可用于检测田间感病葡萄样品中的GLRaV-3。 相似文献
18.
Brian Reavy Maria Sandgren Hugh Barker Pekka Heino Per Oxelfelt 《European journal of plant pathology / European Foundation for Plant Pathology》1997,103(9):829-834
Resistance tests were made on seedlings of transformed lines of Nicotiana benthamiana which contain a transgene encoding the coat protein (CP) gene of a Scottish isolate of potato mop-top virus (PMTV). This transgene has been reported to confer strong resistance to the PMTV isolate from which the transgene sequence was derived and also to a second Scottish isolate. Plants of lines of the transgenic N. benthamiana were as resistant to two Swedish and two Danish PMTV isolates as to a Scottish isolate, and of five lines tested, greater than 93.5% of transgenic plants were immune. The coat protein gene sequences of these four Scandinavian isolates were very similar to those of the two Scottish isolates. The greatest divergence between the isolates was three amino acid changes and there was less than 2% change in CP gene nucleotide sequence. It is concluded that the PMTV CP transgene used in these experiments could confer resistance against isolates from different geographical areas because it is becoming apparent that the CP genes of PMTV isolates are highly conserved. 相似文献
19.
A whitefly transmitted begomovirus was detected by PCR using begomovirus-specific primers from naturally infected Calendula officinalis plants showing yellow vein disease symptoms. An approximately 800 bp PCR amplicon was cloned and sequenced to identify the species of the virus isolate. Analysis of nucleotide sequence data resulted in its identification as the complete coat protein gene open reading frame (CP ORF) of 771 bp, which encoded 256 amino acid residues. The coat protein of the virus isolate shared maximum identities of 96–97% with four strains of Tobacco curly shoot virus (ToCSV) and an Ageratum enation virus (AgEV) during BLAST analysis of sequence data. Nucleotide- and amino acid-based phylogenetic analysis revealed the close relationship of the isolate with ToCSV strains, therefore it has been identified as an isolate of ToCSV and C. officinalis is considered to be a new host of ToCSV begomovirus. Association of a DNA-β molecule with the virus isolate was also detected by PCR and Southern hybridization tests using DNA-β specific primers and probe. 相似文献