Many plant viruses utilize subgenomic RNA as gene expression strategy, therefore mapping subgenomic promoter(SGP) is extremely important for constructing viral vectors. Although Cucumber green mottle mosaic virus(CGMM...Many plant viruses utilize subgenomic RNA as gene expression strategy, therefore mapping subgenomic promoter(SGP) is extremely important for constructing viral vectors. Although Cucumber green mottle mosaic virus(CGMMV)-based virus vectors have been constructed, SGP of the coat protein(CP) has not yet mapped. To this end, we firstly presumed 13 nucleotides upstream of the start codon as the transcription starting site(TSS) as previous study identified by random amplification of c DNA ends(RACE). Secondly, the region from nucleotides –110 to +175 is the putative CP SGP, as predicted, a long stem loop structure by the secondary structure of RNA covering movement protein(MP) and CP. To map the CGMMV CP SGP, we further constructed a series of deletion mutants according to RNA secondary structure prediction. The deletion of TSS upstream significantly enhanced CP transcription when 105 nucleotides were retained before the CP TSS. For the downstream of CP TSS, we analyzed the expression of enhanced green fluorescent protein(EGFP) in a series of vectors with partial deletion of the CGMMV CP and found that the nucleotides from +71 to +91 played a key role in the EGFP expression at the transcription level, while EGFP showed the highest expression level when 160 nucleotides were retained downstream of the CP TSS. To confirm these results, we applied online software MEME to predict the motifs and cis-acting elements in the 466 nucleotides covering the sequences of deletion analysis. Conserved motifs and relative acting elements were in regions in which transcription levels were the highest or enhanced. To our best knowledge, this is the first mapping of CGMMV SGP.展开更多
Cucumber green mottle mosaic virus (CGMMV), a member of the Tobamovirus genus, causes a severe disease of cucurbits. In the Moscow region of Russian Federation, the incidence of infection on cucumber plants in greenho...Cucumber green mottle mosaic virus (CGMMV), a member of the Tobamovirus genus, causes a severe disease of cucurbits. In the Moscow region of Russian Federation, the incidence of infection on cucumber plants in greenhouses is high;however, the virus is poorly studied. In this work, the full-length genomes of two pathogenic MC-1 and MC-2 strains of CGMMV isolated from cucumber plants grown in greenhouses in the Moscow region and the attenuated VIROG-43M strain were sequenced. Comparison of VIROG-43M nucleotide sequence with those of the pathogenic strains revealed three missense mutations. Their role in attenuation is discussed. For the first time, in a number of trials conducted under laboratory conditions and in commercial greenhouses, the efficiency of the attenuated VIROG-43M strain as a biocontrol agent for cucumber plant protection resulting in significant yield gain was demonstrated. Phylogenetic analysis with 83 full-length CGMMV coat protein genes isolated in 16 different countries showed that Russian strains are related to isolates from Spain, Greece, USA and Israel.展开更多
Cucumber green mottle mosaic virus(CGMMV) is a member of the genus Tobamovirus, and is a serious pathogen of Cucurbitaceae crops. Virusderived small interfering RNAs(vsi RNAs), which are processed by Dicer-like and Ar...Cucumber green mottle mosaic virus(CGMMV) is a member of the genus Tobamovirus, and is a serious pathogen of Cucurbitaceae crops. Virusderived small interfering RNAs(vsi RNAs), which are processed by Dicer-like and Argonaute proteins as well as RNA-dependent RNA polymerase,mediate the silencing of viral genomic RNA and host transcripts. To identify the CGMMV derived vsi RNAs and reveal interactions between CGMMV and watermelon host plant, deep sequencing technology was used to identify and characterize the vsi RNAs derived from CGMMV in infected watermelon plants in present study. A total of 10 801 368 vsi RNA reads representing 71 583 unique s RNAs were predicted in CGMMVinoculated watermelon plants. The CGMMV vsi RNAs were mostly 21 or 22 nt long. The majority of the CGMMV vsi RNAs(i.e., 91.7%) originated from the viral sense strand. Additionally, uracil was the predominant 5′-terminal base of vsi RNAs. Furthermore, the putative targets and functions of some of the CGMMV vsi RNAs were predicted and investigated. The results enhance our understanding of the interaction between CGMMV and the host watermelon and provide molecular basis for CGMMV resistance improvement in watermelon and other Cucurbitaceae crops.展开更多
为了揭示黄瓜绿斑驳花叶病毒（Cucumber green mottle mosaic virus,CGMMV）安徽分离物分子变异及系统进化情况,从安徽省5个地区采集感染CGMMV的葫芦科样本。提取感病样本总RNA,经RT-PCR扩增、克隆和测序,获得17个CGMMV分离物的cp基因...为了揭示黄瓜绿斑驳花叶病毒（Cucumber green mottle mosaic virus,CGMMV）安徽分离物分子变异及系统进化情况,从安徽省5个地区采集感染CGMMV的葫芦科样本。提取感病样本总RNA,经RT-PCR扩增、克隆和测序,获得17个CGMMV分离物的cp基因序列。序列比对发现,17个CGMMV安徽分离物的cp基因核苷酸序列相似性极高,达98.4%~100%,与中国及东亚国家和地区的各个CGMMV分离物cp基因核苷酸序列相似性也非常高,达98.6%~100%,而与CGMMV西班牙分离物和俄罗斯分离物cp基因核苷酸序列相似性相对较低,为90.7%~92.6%。从构建的系统关系树可以看出,17个CGMMV安徽分离物与中国及东亚国家和地区的各个CGMMV分离物形成1个单独的分支,而CGMMV俄罗斯与西班牙分离物聚成另1个分支,说明来源于中国和东亚的CGMMV亲缘关系较近。展开更多
对黄瓜绿斑驳花叶病毒(cucumber green mottle mosaic virus,CGMMV)在辽宁省的分布、生物学特性及病毒侵染对西瓜产量和品质的影响进行了研究。结果表明:CGMMV在辽宁省的营口、鞍山、沈阳、丹东、大连、辽阳等地均有分布;在供试的5科21...对黄瓜绿斑驳花叶病毒(cucumber green mottle mosaic virus,CGMMV)在辽宁省的分布、生物学特性及病毒侵染对西瓜产量和品质的影响进行了研究。结果表明:CGMMV在辽宁省的营口、鞍山、沈阳、丹东、大连、辽阳等地均有分布;在供试的5科21种植物中,病毒仅可侵染葫芦科的西瓜、黄瓜、葫芦、南瓜、角瓜、甜瓜、瓠瓜7种植物,并表现系统花叶症状。该病毒致死温度95～100℃,稀释限点10-4～10-5,体外保毒期119d。电镜观察病毒粒子直棒状,长300nm,宽15nm。RT-PCR检测表明,盖州、台安、新民的西瓜花叶样品为黄瓜绿斑驳花叶病毒西瓜株系。利用春秋两季大棚栽培西瓜接种试验表明,西瓜感染CGMMV后,导致植株生长缓慢,结瓜稍小,自根西瓜产量减产幅度很大,秋季大棚减产38.7%～47.6%,春季大棚减产11.4%～19.4%;采用葫芦嫁接的西瓜产量损失变化较小,秋季大棚减产3.4%～17.5%,春季大棚减产2.5%～9.1%。另外,研究结果表明,CGMMV侵染直接导致西瓜倒瓤,使西瓜丧失食用价值,血清学检测倒瓤西瓜携带CGMMV。展开更多
黄瓜绿斑驳花叶病毒(cucumber green mottle mosaic virus,CGMMV)是我国的检疫性有害生物,在广西、辽宁、河北、山东、广东和北京等地均发现疫情,已严重威胁着西瓜、甜瓜、黄瓜等作物的生产,因此加强该病毒的检测极为重要。目前黄瓜绿...黄瓜绿斑驳花叶病毒(cucumber green mottle mosaic virus,CGMMV)是我国的检疫性有害生物,在广西、辽宁、河北、山东、广东和北京等地均发现疫情,已严重威胁着西瓜、甜瓜、黄瓜等作物的生产,因此加强该病毒的检测极为重要。目前黄瓜绿斑驳花叶病毒病的检测主要有生物学检测、血清学检测、分子生物学检测及电镜显微观察等方法。笔者对有关黄瓜绿斑驳花叶病毒病的检测技术进行了综述,并分析了现有检测技术的优缺点和应解决的主要问题,探讨了该病毒检测技术今后发展的方向。展开更多
基金supported by the National Natural Science Foundation of China (31571247)the grants from the earmarked fund for the China Agriculture Research System (CARS-26-13)the Agricultural Science and Technology Innovation Program (ASTIP), Chinese Academy of Agricultural Sciences (CAAS-ASTIP-2018-ZFRI-08)
文摘Many plant viruses utilize subgenomic RNA as gene expression strategy, therefore mapping subgenomic promoter(SGP) is extremely important for constructing viral vectors. Although Cucumber green mottle mosaic virus(CGMMV)-based virus vectors have been constructed, SGP of the coat protein(CP) has not yet mapped. To this end, we firstly presumed 13 nucleotides upstream of the start codon as the transcription starting site(TSS) as previous study identified by random amplification of c DNA ends(RACE). Secondly, the region from nucleotides –110 to +175 is the putative CP SGP, as predicted, a long stem loop structure by the secondary structure of RNA covering movement protein(MP) and CP. To map the CGMMV CP SGP, we further constructed a series of deletion mutants according to RNA secondary structure prediction. The deletion of TSS upstream significantly enhanced CP transcription when 105 nucleotides were retained before the CP TSS. For the downstream of CP TSS, we analyzed the expression of enhanced green fluorescent protein(EGFP) in a series of vectors with partial deletion of the CGMMV CP and found that the nucleotides from +71 to +91 played a key role in the EGFP expression at the transcription level, while EGFP showed the highest expression level when 160 nucleotides were retained downstream of the CP TSS. To confirm these results, we applied online software MEME to predict the motifs and cis-acting elements in the 466 nucleotides covering the sequences of deletion analysis. Conserved motifs and relative acting elements were in regions in which transcription levels were the highest or enhanced. To our best knowledge, this is the first mapping of CGMMV SGP.
文摘Cucumber green mottle mosaic virus (CGMMV), a member of the Tobamovirus genus, causes a severe disease of cucurbits. In the Moscow region of Russian Federation, the incidence of infection on cucumber plants in greenhouses is high;however, the virus is poorly studied. In this work, the full-length genomes of two pathogenic MC-1 and MC-2 strains of CGMMV isolated from cucumber plants grown in greenhouses in the Moscow region and the attenuated VIROG-43M strain were sequenced. Comparison of VIROG-43M nucleotide sequence with those of the pathogenic strains revealed three missense mutations. Their role in attenuation is discussed. For the first time, in a number of trials conducted under laboratory conditions and in commercial greenhouses, the efficiency of the attenuated VIROG-43M strain as a biocontrol agent for cucumber plant protection resulting in significant yield gain was demonstrated. Phylogenetic analysis with 83 full-length CGMMV coat protein genes isolated in 16 different countries showed that Russian strains are related to isolates from Spain, Greece, USA and Israel.
基金Project funded by China Postdoctoral Science Foundation (2016M601973)the National Natural Science Foundation of China (31572145 and 31272188)Zhejiang Major Agricultural Science and Technology Projects for New Varieties Breeding (2016C02051-4-2)
文摘Cucumber green mottle mosaic virus(CGMMV) is a member of the genus Tobamovirus, and is a serious pathogen of Cucurbitaceae crops. Virusderived small interfering RNAs(vsi RNAs), which are processed by Dicer-like and Argonaute proteins as well as RNA-dependent RNA polymerase,mediate the silencing of viral genomic RNA and host transcripts. To identify the CGMMV derived vsi RNAs and reveal interactions between CGMMV and watermelon host plant, deep sequencing technology was used to identify and characterize the vsi RNAs derived from CGMMV in infected watermelon plants in present study. A total of 10 801 368 vsi RNA reads representing 71 583 unique s RNAs were predicted in CGMMVinoculated watermelon plants. The CGMMV vsi RNAs were mostly 21 or 22 nt long. The majority of the CGMMV vsi RNAs(i.e., 91.7%) originated from the viral sense strand. Additionally, uracil was the predominant 5′-terminal base of vsi RNAs. Furthermore, the putative targets and functions of some of the CGMMV vsi RNAs were predicted and investigated. The results enhance our understanding of the interaction between CGMMV and the host watermelon and provide molecular basis for CGMMV resistance improvement in watermelon and other Cucurbitaceae crops.
文摘为了揭示黄瓜绿斑驳花叶病毒（Cucumber green mottle mosaic virus,CGMMV）安徽分离物分子变异及系统进化情况,从安徽省5个地区采集感染CGMMV的葫芦科样本。提取感病样本总RNA,经RT-PCR扩增、克隆和测序,获得17个CGMMV分离物的cp基因序列。序列比对发现,17个CGMMV安徽分离物的cp基因核苷酸序列相似性极高,达98.4%~100%,与中国及东亚国家和地区的各个CGMMV分离物cp基因核苷酸序列相似性也非常高,达98.6%~100%,而与CGMMV西班牙分离物和俄罗斯分离物cp基因核苷酸序列相似性相对较低,为90.7%~92.6%。从构建的系统关系树可以看出,17个CGMMV安徽分离物与中国及东亚国家和地区的各个CGMMV分离物形成1个单独的分支,而CGMMV俄罗斯与西班牙分离物聚成另1个分支,说明来源于中国和东亚的CGMMV亲缘关系较近。
文摘对黄瓜绿斑驳花叶病毒(cucumber green mottle mosaic virus,CGMMV)在辽宁省的分布、生物学特性及病毒侵染对西瓜产量和品质的影响进行了研究。结果表明:CGMMV在辽宁省的营口、鞍山、沈阳、丹东、大连、辽阳等地均有分布;在供试的5科21种植物中,病毒仅可侵染葫芦科的西瓜、黄瓜、葫芦、南瓜、角瓜、甜瓜、瓠瓜7种植物,并表现系统花叶症状。该病毒致死温度95～100℃,稀释限点10-4～10-5,体外保毒期119d。电镜观察病毒粒子直棒状,长300nm,宽15nm。RT-PCR检测表明,盖州、台安、新民的西瓜花叶样品为黄瓜绿斑驳花叶病毒西瓜株系。利用春秋两季大棚栽培西瓜接种试验表明,西瓜感染CGMMV后,导致植株生长缓慢,结瓜稍小,自根西瓜产量减产幅度很大,秋季大棚减产38.7%～47.6%,春季大棚减产11.4%～19.4%;采用葫芦嫁接的西瓜产量损失变化较小,秋季大棚减产3.4%～17.5%,春季大棚减产2.5%～9.1%。另外,研究结果表明,CGMMV侵染直接导致西瓜倒瓤,使西瓜丧失食用价值,血清学检测倒瓤西瓜携带CGMMV。
文摘黄瓜绿斑驳花叶病毒(cucumber green mottle mosaic virus,CGMMV)是我国的检疫性有害生物,在广西、辽宁、河北、山东、广东和北京等地均发现疫情,已严重威胁着西瓜、甜瓜、黄瓜等作物的生产,因此加强该病毒的检测极为重要。目前黄瓜绿斑驳花叶病毒病的检测主要有生物学检测、血清学检测、分子生物学检测及电镜显微观察等方法。笔者对有关黄瓜绿斑驳花叶病毒病的检测技术进行了综述,并分析了现有检测技术的优缺点和应解决的主要问题,探讨了该病毒检测技术今后发展的方向。
