柑橘碎叶病毒侵染性克隆构建及其诱导的基因沉默（VIGS）体系建立
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摘要
柑橘是世界第一大水果。我国柑橘栽培面积超过290万公顷,产量达3000万吨,均居世界第一位。随着克里曼丁红橘(Citrus clementina Hort. ex Tan)和甜橙(C. sinensis cv. Valencia)全基因组序列的公布,以及柑橘抗病、抗逆等相关基因序列表达标签(Expressed sequence tags, EST)的大量累积,柑橘基因功能解析已成为目前的研究热点和挑战之一。病毒诱导的基因沉默(Virus-induced gene silencing, VIGS)是近年来发展的一种基因功能研究工具,与转基因、基因敲除、反义抑制等常用生物技术相比,试验周期短,不需要遗传转化,具有操作简便、成本低、高通量等优势。利用VIGS技术有望克服柑橘这种多年生木本作物童期长、遗传转化困难等瓶颈,加速其基因功能研究的进展。为此,本研究在柑橘碎叶病毒(Citrus tatter leaf virus, CTLV)检测方法及多态性研究的基础上,筛选CTLV分离株进行全基因组cDNA扩增、侵染性克隆构建,并进一步开展了在模式植物本生烟(Nicotiana benthamiana)上建立VIGS体系的研究,以期为在柑橘上实施VIGS进而研究其基因功能奠定基础。主要研究结果如下：
1. CTLV一步法及巢式RT-PCR检测体系建立
通过引物比对,建立了CTLV的一步法及巢式RT-PCR检测体系。其中,一步法RT-PCR产物889bp,包含整个CP及3'UTR序列,比目前的常规RT-PCR操作简便,污染几率小；巢式RT-PCR检测灵敏度高,比一步法RT-PCR至少提高100倍,检测模板总核酸的最低浓度约1.27pg/μL。
2. CTLV多态性分析
建立了CTLV分子变异的Hinf Ⅰ/RFLP检测体系,定义RFLP组群9个,发现CTLV以单一RFLP组群侵染为主,其中RFLP Ⅰ和RFLP Ⅱ组群可能是优势流行组群,但也存在混合侵染。该体系简便、快速、重复性好,可用于大规模样品的分子变异检测。
对收集保存的18份CTLV分离株进行了指示植物鉴定、3’端序列分析及RFLP检测。结果表明,在指示植物腊斯克枳橙(C. sinensis×Poncirus trifoliata cv. Rusk)上表现弱到中等强度症状的6个分离株均属于或携带有RFLP Ⅱ或RFLPⅢ组群,而其它RFLP组群均表现出强毒株特征。在根据3'端序列(889bp)构建的系统进化树上,CTLV分离株聚为了2个同源性组群,同源性组群A包含的8个序列均为RFLP Ⅱ或RFLP Ⅲ组群；同源性组群B则包含了其它RFLP组群的所有序列。同时,在指示植物上表现较弱症状的多数(4/6)CTLV分离物划分在A组群,多数(10/12)CTLV强毒分离物则划分在B组群。
可见,指示植物上的弱(中)毒分离株侵染症状-RFLPⅡ或RFLPⅢ组群-同源性A组群三者之间可能存在某种内在联系。RFLP组群检测可用于研究CTLV的分子变异,对于快速鉴定CTLV致病性强弱可能具有参考价值。
3. CTLV基因组全长cDNA扩增及序列分析
在通过RACE技术获得末端精确序列的基础上,建立了CTLV全长的RT-PCR体系,进而扩增并克隆了CTLV新会橙分离株及满头红分离株基因组全长cDNA。序列分析表明,CTLV-XHC(登录号KC588947)和CTLV-MTH(登录号KC588948)基因组全长均为6497nt[不包括3'poly(A)尾巴],均包含2个开放读码框,在推定的CP及MP上游均具有保守的亚基因组启动子核心序列‘'UUAGGU",与目前已报道的CTLV及苹果茎沟病毒(Apple stem grooving virus, ASGV)的基因组结构一致。
同登录GenBank的所有CTLV及ASGV全基因组序列比对显示,CTLV-XHC与我国砂糖橘分离株CTLV-S序列一致性最高,为98%,在依据全基因组核苷酸序列构建的系统进化树上,CTLV-XHC与柑橘来源的CTLV分离株聚为一簇；有趣的是,CTLV-MTH与日本ASGV分离物序列一致性最高,为91%,而与台湾甜橙分离株CTLV-LC一致性最低,为82.0%,在系统进化树上CTLV-MTH与多数ASGV分离株聚为一簇。这一结果对于CTLV与ASGV的起源进化关系具有启示意义。
4. CTLV侵染性克隆构建
对6个CTLV基因组全长cDNA克隆进行体外转录并摩擦接种昆诺藜(Chenopodium quinoa),症状观察及RT-PCR检测结果表明,CTLV-XHC和CTLV-MTH的体外转录物具有侵染性。
为获得适于农杆菌介导接种的CTLV侵染性克隆,以pCAMBIA1301为骨架构建了双元植物表达载体pCSSN:LB-2x35S-MCS-NOS-RB,通过瞬时表达GFP验证其有效性后,采用分步克隆策略将CTLV-XHC和CTLV-MTH分别插入到pCSSN载体2x35S启动子与NOS终止子之间。经筛选、鉴定获得了CTLV侵染性克隆pCSSN-XHC和pCSSN-MTH。农杆菌介导的本生烟(N. benthamiana)注射接种实验表明,二者的侵染症状与野生型病毒对照无明显差异,接种时共表达沉默抑制子p19提高了pCSSN-XHC和pCSSN-MTH的侵染率。
5. CTLV诱导的VIGS体系建立
以CTLV侵染性克隆pCSSN-XHC为基础,通过重组克隆在CP终止子之后引入NruⅠ识别序列,构建了VIGS载体pCTLV-00:LB-2x35S-XHC(Nru Ⅰ)-NOS-RB。反向插入402bp源于本生烟的八氢番茄红素脱氢酶(Phytoene desaturase, PDS)基因片段后,重组载体pCTLV-PDS402R通过农杆菌浸润接种本生烟,在22℃光照16h和20℃黑暗8h周期条件下,诱导产生了PDS基因沉默的白化表型。Real-time RT-PCR检测结果表明,pCTLV-PDS402R侵染植株PDS基因的相对表达量比空载体pCTLV-00对照下降约25%。说明基于CTLV上的VIGS体系初步建立。
为提高VIGS沉默效率,引入具有自我剪切功能的核酶序列到pCTLV-00,构建了VIGS载体pCTLV1:LB-2x35S-XHC(Nru Ⅰ)-RZ-NOS-RB;进一步分别引入135bp、105bp、90bp和61bp的CP亚基因组启动子片段构建了VIGS载体pCTLV2、pCTLV3、 pCTLV4、pCTLV5。上述系列VIGS载体均反向插入了PDS基因片段,基因沉默效应正在评估当中。
Citrus is the first important fruit crop by total output in the world. China ranks No.1with a total acreage of over2.9million hectares and an annual citrus production of over30million tons. At present, the genome sequence of clemantine tangor(Citrus clementina Hort. ex Tan and valencia sweet orange(C. sinensis cv. Valencia) have been published, and a large number of expressed sequence tags (ESTs) has been released to GenBank. In contrast to the enormous progress in obtaining sequence data, the identification of citrus genes functions has been being focused with challenge by its long juvenile period and genetic transformation difficulty. A tool to bridge the gap between sequence information and gene functions for citrus is in demand. Virus-induced gene silencing (VIGS) is a recently developed tool for gene function studies, which is particularly attractive for such woody species as citrus. Compared to commonly used biotechnological tools such as transgenic mean, gene deletion method and inhibition technique by antisense nucleotides, VIGS is rapid with low cost, no need for stable plant transformation, and allows a large-scale screening of genes for functional analysis. The aim of this paper was to develop viral vectors for gene silencing that might be useful to identify gene functions and for citrus genetic improvement. For this purpose, Citrus tatter leaf virus (CTLV) isolates were collected and detected by newly established One-step or Nested RT-PCR, and then were addressed to biological indexing and genetic polymorphism assay. Selected isolates were adopted to construct full-length cDNA clone of CTLV genome, followed by sequencing and engineering to a modified binary vector. Based on the clone shown to be infectious, a series VIGS vectors were developed and evaluated by silencing phytoene desaturase gene (PDS) upon agroinoculation into model plant leaves of tabacco Nicotiana benthamiana. The outputs are as follows.
1. One-step and Nested RT-PCR for CTLV detection
A one-step RT-PCR and a nested RT-PCR were established for CTLV detection by comparing primer pairs. With a product of889bp in length, the One-step RT-PCR was more convenient and lower risk of contamination than that of conventional RT-PCR. The Nested RT-PCR was at least hundred times more sensitive than that of the One-step RT-PCR, and the lowest detection limit of the method reached1.27pg/μL, which could be very useful for CTLV diagnostic and certification programs.
2. Genetic diversity of CTLV
A method for detecting molecular variation of CTLV was developed based on restriction fragment length polymorphisms (RFLP) of the3'sequence amplified by RT-PCR, and nine Hinf I RFLP patterns were defined. The method was rapid, reliable, reproducible, and could be used for a large scale of samples. Detection data indicated that the majority of CTLV isolates presented as single well-defined patterns, among which RFLP I and RFLP Ⅱ were dominant. In contrast, a few samples presented as a mixture of two RFLP patterns, suggesting a mixed infection of CTLV variants.
Eighteen isolates of CTLV were characterized by biological indexing, sequencing and RFLP assay. Six isolates induced mild to moderate symptoms on indexing plants (C. sinensis×Poncirus trifoliata cv. Rusk), and all of them were conformed to carry with RFLP II or RFLP Ⅲ. In contrast, four isolates carrying with RFLP I and six other samples exhibited RFLPⅣ-Ⅸ induced severe symptoms. In the phylogenetic tree constructed according to3' nucleotide sequence, CTLV isolates tested were divided into two clusters clearly, of which phylogenetic groups A, included all sequence were conformed to carry with RFLP II or RFLP III patterns, while phylogenetic groups B included other RFLP restrictotypes. Also, phylogenetic groups A included the majority mild (moderate) strain (4/6), while phylogenetic groups B included the majority severe strain (10/12).
The results above revealed somewhat correlation among mild (moderate) symptoms, phylogenetic groups A and RFLP Ⅱ or RFLP Ⅲ restrictotypes, suggesting RFLP analysis may be useful in quick identification of mild strains of CTLV.
3. RT-PCR amplification and Sequence analyses of CTLV
A single step RT-PCR for detecting the entire CTLV genomic RNA were established with a pair of primers specific to its exact5'and3'ends sequences, which were determined by RACE system (Clontech). Full-length cDNA of CTLV isolates XHC and MTH were successfully amplified according to the method, and then were cloned and sequenced. The complete genome sequence of CTLV-XHC and CTLV-MTH were determined to be6497nucleotides in length, excluding the3'-terminal poly(A) tract, and contained two putative overlapping open reading frames (ORFs). For the genome of CTLV-XHC and CTLV-MTH, the core sequences of the sgRNA promoter, UUAGGU, were also found upstream from the transcription start sites of both the putative CP and MP sgRNAs, which is broadly conserved among viruses in the Flexiviridae.
Homology matrix of nucleotide sequences of CTLV-XHC, CTLV-MTH and other available Capilloviruses in GenBank was conducted, and then the phylogenetic tree derived from those sequences was constructed. CTLV-XHC has the maximum identity (98%) with CTLV-S, a strain isolated from Shatangju(C. reticulata) in China, and clustered together with isolates from citrus in the phylogenetic tree. Interestingly, CTLV-MTH showed the highest identity (91%) with an ASGV isolate from Japan, and the lowest identity (82.0%) with a CTLV isolate from sweet orange located at Taiwan. In the phylogenetic tree, CTLV-MTH was clustered together with most of ASGV isolates. These results may have enlightenment significance for CTLV and ASGV evolutionary relationships.
4. Construction of full-length infectious clones of CTLV
Capped in vitro transcripts of six full-length clones of CTLV were generated, and inoculated mechanically onto leaves of Chenopodium quinoa plants, and evaluated periodically for symptom development. In vitro transcripts from CTLV-XHC and CTLV-MTH elicited systemic infections in some of the inoculated plants, which were confirmed both by RT-PCR detection and symptom observation.
In order to generate the agrobacterium-mediated infectious cDNA clones of CTLV, a binary vector pCSSN (LB-2x35S-MCS-NOS-RB) was firstly constructed based on pCAMBIA1301backbone, and evaluated for validity by GFP expression. The full-length cDNAs of CTLV-XHC and CTLV-MTH were inserted into pCSSN between2x35S promoter and NOS terminator respectively, and then were introduced into agrobacterium EHA105and inoculated N. benthamiana by agro-infiltration with or without co-expressing of the p19silencing suppressor. As a result, pCSSN-XHC and pCSSN-MTH were obtained with the corresponding full-length cDNA of CTLV via duplicated CaMV35S promoter, which were confirmed by sequencing. Part of plants agro-infiltrated with pCSSN-XHC or pCSSN-MTH were infected successfully according to RT-PCR results, and developed symptoms similar to those induced by the wild-type CTLV, but with a3-5d delay. When the p19silencing suppressor involved in the process of agrobacterium-mediated inoculation, higher infectivity of pCSSN-XHC or pCSSN-MTH were observed.
5. Developing VIGS vector based on CTLV
A VIGS vector pCTLV-00[LB-2x35S-XHC(Nru I)-NOS-RB] was constructed by introducing a unique Nru I restriction site to pCSSN-XHC at closely downstream of the CP. A402bp fragment of PDS from N. benthamiana was RT-PCR amplified and inserted into the restriction site in the antisense orientation. The recombinant (pCTLV-PDS402R) was inoculated onto leaves of N. benthamiana by agro-infiltration with co-expressing the p19silencing suppressor. Grown in a plant growth chamber at22/20℃day/night, and16/8h light/dark regime, some of those plants initiated white spots on the upper leaves25-30d post inoculation, indicating the PDS had been silenced. Quantification of PDS-mRNA by Real-time RT-PCR showed that tobacco leaves with a silencing phenotype had25%lower mRNA accumulation than that of leaves from plants infected with pCTLV-00. These results suggest that a VIGS system based on CTLV was preliminarily established.
In order to improve effciency of the CTLV-mediated VIGS system, pCTLV1[LB-2x35S-XHC(Nru I)-RZ-NOS-RB] was developed by adding a sequence of Hepatitis delta virus ribozyme to pCTLV-00before its NOS terminator. Further modifications were done based on pCTLV1to generate pCTLV2, pCTLV3, pCTLV4, and pCTLV5, which contained a duplicated fragment of CP sgRNA promoter in different lengths(135bp,105bp,90bp, and61bp respectively). These vectors had been inserted with PDS fragment, and their efficiency were evaluating.
1. CTLV一步法及巢式RT-PCR检测体系建立
通过引物比对,建立了CTLV的一步法及巢式RT-PCR检测体系。其中,一步法RT-PCR产物889bp,包含整个CP及3'UTR序列,比目前的常规RT-PCR操作简便,污染几率小；巢式RT-PCR检测灵敏度高,比一步法RT-PCR至少提高100倍,检测模板总核酸的最低浓度约1.27pg/μL。
2. CTLV多态性分析
建立了CTLV分子变异的Hinf Ⅰ/RFLP检测体系,定义RFLP组群9个,发现CTLV以单一RFLP组群侵染为主,其中RFLP Ⅰ和RFLP Ⅱ组群可能是优势流行组群,但也存在混合侵染。该体系简便、快速、重复性好,可用于大规模样品的分子变异检测。
对收集保存的18份CTLV分离株进行了指示植物鉴定、3’端序列分析及RFLP检测。结果表明,在指示植物腊斯克枳橙(C. sinensis×Poncirus trifoliata cv. Rusk)上表现弱到中等强度症状的6个分离株均属于或携带有RFLP Ⅱ或RFLPⅢ组群,而其它RFLP组群均表现出强毒株特征。在根据3'端序列(889bp)构建的系统进化树上,CTLV分离株聚为了2个同源性组群,同源性组群A包含的8个序列均为RFLP Ⅱ或RFLP Ⅲ组群；同源性组群B则包含了其它RFLP组群的所有序列。同时,在指示植物上表现较弱症状的多数(4/6)CTLV分离物划分在A组群,多数(10/12)CTLV强毒分离物则划分在B组群。
可见,指示植物上的弱(中)毒分离株侵染症状-RFLPⅡ或RFLPⅢ组群-同源性A组群三者之间可能存在某种内在联系。RFLP组群检测可用于研究CTLV的分子变异,对于快速鉴定CTLV致病性强弱可能具有参考价值。
3. CTLV基因组全长cDNA扩增及序列分析
在通过RACE技术获得末端精确序列的基础上,建立了CTLV全长的RT-PCR体系,进而扩增并克隆了CTLV新会橙分离株及满头红分离株基因组全长cDNA。序列分析表明,CTLV-XHC(登录号KC588947)和CTLV-MTH(登录号KC588948)基因组全长均为6497nt[不包括3'poly(A)尾巴],均包含2个开放读码框,在推定的CP及MP上游均具有保守的亚基因组启动子核心序列‘'UUAGGU",与目前已报道的CTLV及苹果茎沟病毒(Apple stem grooving virus, ASGV)的基因组结构一致。
同登录GenBank的所有CTLV及ASGV全基因组序列比对显示,CTLV-XHC与我国砂糖橘分离株CTLV-S序列一致性最高,为98%,在依据全基因组核苷酸序列构建的系统进化树上,CTLV-XHC与柑橘来源的CTLV分离株聚为一簇；有趣的是,CTLV-MTH与日本ASGV分离物序列一致性最高,为91%,而与台湾甜橙分离株CTLV-LC一致性最低,为82.0%,在系统进化树上CTLV-MTH与多数ASGV分离株聚为一簇。这一结果对于CTLV与ASGV的起源进化关系具有启示意义。
4. CTLV侵染性克隆构建
对6个CTLV基因组全长cDNA克隆进行体外转录并摩擦接种昆诺藜(Chenopodium quinoa),症状观察及RT-PCR检测结果表明,CTLV-XHC和CTLV-MTH的体外转录物具有侵染性。
为获得适于农杆菌介导接种的CTLV侵染性克隆,以pCAMBIA1301为骨架构建了双元植物表达载体pCSSN:LB-2x35S-MCS-NOS-RB,通过瞬时表达GFP验证其有效性后,采用分步克隆策略将CTLV-XHC和CTLV-MTH分别插入到pCSSN载体2x35S启动子与NOS终止子之间。经筛选、鉴定获得了CTLV侵染性克隆pCSSN-XHC和pCSSN-MTH。农杆菌介导的本生烟(N. benthamiana)注射接种实验表明,二者的侵染症状与野生型病毒对照无明显差异,接种时共表达沉默抑制子p19提高了pCSSN-XHC和pCSSN-MTH的侵染率。
5. CTLV诱导的VIGS体系建立
以CTLV侵染性克隆pCSSN-XHC为基础,通过重组克隆在CP终止子之后引入NruⅠ识别序列,构建了VIGS载体pCTLV-00:LB-2x35S-XHC(Nru Ⅰ)-NOS-RB。反向插入402bp源于本生烟的八氢番茄红素脱氢酶(Phytoene desaturase, PDS)基因片段后,重组载体pCTLV-PDS402R通过农杆菌浸润接种本生烟,在22℃光照16h和20℃黑暗8h周期条件下,诱导产生了PDS基因沉默的白化表型。Real-time RT-PCR检测结果表明,pCTLV-PDS402R侵染植株PDS基因的相对表达量比空载体pCTLV-00对照下降约25%。说明基于CTLV上的VIGS体系初步建立。
为提高VIGS沉默效率,引入具有自我剪切功能的核酶序列到pCTLV-00,构建了VIGS载体pCTLV1:LB-2x35S-XHC(Nru Ⅰ)-RZ-NOS-RB;进一步分别引入135bp、105bp、90bp和61bp的CP亚基因组启动子片段构建了VIGS载体pCTLV2、pCTLV3、 pCTLV4、pCTLV5。上述系列VIGS载体均反向插入了PDS基因片段,基因沉默效应正在评估当中。
Citrus is the first important fruit crop by total output in the world. China ranks No.1with a total acreage of over2.9million hectares and an annual citrus production of over30million tons. At present, the genome sequence of clemantine tangor(Citrus clementina Hort. ex Tan and valencia sweet orange(C. sinensis cv. Valencia) have been published, and a large number of expressed sequence tags (ESTs) has been released to GenBank. In contrast to the enormous progress in obtaining sequence data, the identification of citrus genes functions has been being focused with challenge by its long juvenile period and genetic transformation difficulty. A tool to bridge the gap between sequence information and gene functions for citrus is in demand. Virus-induced gene silencing (VIGS) is a recently developed tool for gene function studies, which is particularly attractive for such woody species as citrus. Compared to commonly used biotechnological tools such as transgenic mean, gene deletion method and inhibition technique by antisense nucleotides, VIGS is rapid with low cost, no need for stable plant transformation, and allows a large-scale screening of genes for functional analysis. The aim of this paper was to develop viral vectors for gene silencing that might be useful to identify gene functions and for citrus genetic improvement. For this purpose, Citrus tatter leaf virus (CTLV) isolates were collected and detected by newly established One-step or Nested RT-PCR, and then were addressed to biological indexing and genetic polymorphism assay. Selected isolates were adopted to construct full-length cDNA clone of CTLV genome, followed by sequencing and engineering to a modified binary vector. Based on the clone shown to be infectious, a series VIGS vectors were developed and evaluated by silencing phytoene desaturase gene (PDS) upon agroinoculation into model plant leaves of tabacco Nicotiana benthamiana. The outputs are as follows.
1. One-step and Nested RT-PCR for CTLV detection
A one-step RT-PCR and a nested RT-PCR were established for CTLV detection by comparing primer pairs. With a product of889bp in length, the One-step RT-PCR was more convenient and lower risk of contamination than that of conventional RT-PCR. The Nested RT-PCR was at least hundred times more sensitive than that of the One-step RT-PCR, and the lowest detection limit of the method reached1.27pg/μL, which could be very useful for CTLV diagnostic and certification programs.
2. Genetic diversity of CTLV
A method for detecting molecular variation of CTLV was developed based on restriction fragment length polymorphisms (RFLP) of the3'sequence amplified by RT-PCR, and nine Hinf I RFLP patterns were defined. The method was rapid, reliable, reproducible, and could be used for a large scale of samples. Detection data indicated that the majority of CTLV isolates presented as single well-defined patterns, among which RFLP I and RFLP Ⅱ were dominant. In contrast, a few samples presented as a mixture of two RFLP patterns, suggesting a mixed infection of CTLV variants.
Eighteen isolates of CTLV were characterized by biological indexing, sequencing and RFLP assay. Six isolates induced mild to moderate symptoms on indexing plants (C. sinensis×Poncirus trifoliata cv. Rusk), and all of them were conformed to carry with RFLP II or RFLP Ⅲ. In contrast, four isolates carrying with RFLP I and six other samples exhibited RFLPⅣ-Ⅸ induced severe symptoms. In the phylogenetic tree constructed according to3' nucleotide sequence, CTLV isolates tested were divided into two clusters clearly, of which phylogenetic groups A, included all sequence were conformed to carry with RFLP II or RFLP III patterns, while phylogenetic groups B included other RFLP restrictotypes. Also, phylogenetic groups A included the majority mild (moderate) strain (4/6), while phylogenetic groups B included the majority severe strain (10/12).
The results above revealed somewhat correlation among mild (moderate) symptoms, phylogenetic groups A and RFLP Ⅱ or RFLP Ⅲ restrictotypes, suggesting RFLP analysis may be useful in quick identification of mild strains of CTLV.
3. RT-PCR amplification and Sequence analyses of CTLV
A single step RT-PCR for detecting the entire CTLV genomic RNA were established with a pair of primers specific to its exact5'and3'ends sequences, which were determined by RACE system (Clontech). Full-length cDNA of CTLV isolates XHC and MTH were successfully amplified according to the method, and then were cloned and sequenced. The complete genome sequence of CTLV-XHC and CTLV-MTH were determined to be6497nucleotides in length, excluding the3'-terminal poly(A) tract, and contained two putative overlapping open reading frames (ORFs). For the genome of CTLV-XHC and CTLV-MTH, the core sequences of the sgRNA promoter, UUAGGU, were also found upstream from the transcription start sites of both the putative CP and MP sgRNAs, which is broadly conserved among viruses in the Flexiviridae.
Homology matrix of nucleotide sequences of CTLV-XHC, CTLV-MTH and other available Capilloviruses in GenBank was conducted, and then the phylogenetic tree derived from those sequences was constructed. CTLV-XHC has the maximum identity (98%) with CTLV-S, a strain isolated from Shatangju(C. reticulata) in China, and clustered together with isolates from citrus in the phylogenetic tree. Interestingly, CTLV-MTH showed the highest identity (91%) with an ASGV isolate from Japan, and the lowest identity (82.0%) with a CTLV isolate from sweet orange located at Taiwan. In the phylogenetic tree, CTLV-MTH was clustered together with most of ASGV isolates. These results may have enlightenment significance for CTLV and ASGV evolutionary relationships.
4. Construction of full-length infectious clones of CTLV
Capped in vitro transcripts of six full-length clones of CTLV were generated, and inoculated mechanically onto leaves of Chenopodium quinoa plants, and evaluated periodically for symptom development. In vitro transcripts from CTLV-XHC and CTLV-MTH elicited systemic infections in some of the inoculated plants, which were confirmed both by RT-PCR detection and symptom observation.
In order to generate the agrobacterium-mediated infectious cDNA clones of CTLV, a binary vector pCSSN (LB-2x35S-MCS-NOS-RB) was firstly constructed based on pCAMBIA1301backbone, and evaluated for validity by GFP expression. The full-length cDNAs of CTLV-XHC and CTLV-MTH were inserted into pCSSN between2x35S promoter and NOS terminator respectively, and then were introduced into agrobacterium EHA105and inoculated N. benthamiana by agro-infiltration with or without co-expressing of the p19silencing suppressor. As a result, pCSSN-XHC and pCSSN-MTH were obtained with the corresponding full-length cDNA of CTLV via duplicated CaMV35S promoter, which were confirmed by sequencing. Part of plants agro-infiltrated with pCSSN-XHC or pCSSN-MTH were infected successfully according to RT-PCR results, and developed symptoms similar to those induced by the wild-type CTLV, but with a3-5d delay. When the p19silencing suppressor involved in the process of agrobacterium-mediated inoculation, higher infectivity of pCSSN-XHC or pCSSN-MTH were observed.
5. Developing VIGS vector based on CTLV
A VIGS vector pCTLV-00[LB-2x35S-XHC(Nru I)-NOS-RB] was constructed by introducing a unique Nru I restriction site to pCSSN-XHC at closely downstream of the CP. A402bp fragment of PDS from N. benthamiana was RT-PCR amplified and inserted into the restriction site in the antisense orientation. The recombinant (pCTLV-PDS402R) was inoculated onto leaves of N. benthamiana by agro-infiltration with co-expressing the p19silencing suppressor. Grown in a plant growth chamber at22/20℃day/night, and16/8h light/dark regime, some of those plants initiated white spots on the upper leaves25-30d post inoculation, indicating the PDS had been silenced. Quantification of PDS-mRNA by Real-time RT-PCR showed that tobacco leaves with a silencing phenotype had25%lower mRNA accumulation than that of leaves from plants infected with pCTLV-00. These results suggest that a VIGS system based on CTLV was preliminarily established.
In order to improve effciency of the CTLV-mediated VIGS system, pCTLV1[LB-2x35S-XHC(Nru I)-RZ-NOS-RB] was developed by adding a sequence of Hepatitis delta virus ribozyme to pCTLV-00before its NOS terminator. Further modifications were done based on pCTLV1to generate pCTLV2, pCTLV3, pCTLV4, and pCTLV5, which contained a duplicated fragment of CP sgRNA promoter in different lengths(135bp,105bp,90bp, and61bp respectively). These vectors had been inserted with PDS fragment, and their efficiency were evaluating.
引文
[1]Miyakawa T., Ito T. Tatter leaf-citrange stunt [M], Compendium of citrus diseases.2000, APS Press, St. Paul, MN. p60.
[2]Wallace J. M., Drake R. J. Tatter leaf, a previously undesoribed virus effect on citrus [J]. Plant Disease Reporter,1962,46(4):211-212.
[3]Yarwood C. E. Mechanical transmission of a latent virus [J]. Phytopathology, 1963,53:1145.
[4]Fulton R. W. Mechanical transmission of tatter leaf virus from cowpea to citrus [J]. Phytopathology,1966,56(5):575.
[5]Miyakawa T., Matsui C. A bud-union abnormality of satsuma mandarin on Poncirus trifoliata rootstock in Japan [C]. Proceedings of the 7th Conference of the International Organization of Citrus Virologists. Riverside:IOCV, 1976:125-131.
[6]Miyakawa T., Tsuji M. The association of tatter leaf virus with bud-union crease of trees on trifoliate orange rootstock [C]. Proceedings of the 10th Conference of the International Organization of Citrus Virologists. Riverside:IOCV, 1988:360-364.
[7]Broadbent P., Dephoff C. M., Gilkeson C. Detection of Citrus tatter leaf virus in Australia [J]. Australasian Plant Pathology,1994,23(1):20-24.
[8]Da Graca J. V. Citrus tatter leaf virus in South African Meyer lemon [J]. Citrus Subtrop. Fruit J.,1977,529:18.
[9]Zhang T. M., Liang X. Y., Roistacher C. N. Occurrence and detection of Citrus tatter leaf virus (CTLV) in Huangyan, Zhejiang province, China [J]. Plant Disease,1988,72:543-545.
[10]Fajinmi A. A, Fajinmi O. B, Amusa N. A. An overview of citrus virus disease and its control in Nigeria [J]. Journal of Advances in Developmental Research,2011, 2(2):151-157.
[11]刘干生,刘震,刘尚泉等.柑橘碎叶病危害的调查[J].中国南方果树,2003,32(2)：22.
[12]赵小龙,张社南,刘升球等.柑橘碎叶病及其防治[J].广西园艺,2005,16(2)：32-33.
[13]Chung K., Jian W. R. Occurrence and distribution of citrus tatter leaf in Fujian, China [J].1990.
[14]Schwarz R. E. Mechanical transmission of a virus from Meyer lemon to herbaceous hosts [J]. South African Journal of Agriculture Science,1966,9: 263-264.
[15]Wallace J. M., Drake R. J. New information on symptom effects and host range of the Citrus tatter-leaf virus [J]. Plant Disease Reporter,1963,47:352-353.
[16]Roistacher C. N. A historical review of the major graft-transmissible diseases of citrus [M]. Roma (Italia):Food and Agriculture Organization of the United Nations, Regional Office for the Near East,1995.
[17]Roistacher C. N. Citrus tatter leaf virus:Further evidence for single virus complex [C]. Proceedings of the 10th Conference of the International Organization of Citrus Virologists. Riverside:IOCV,1988:353-359.
[18]Iwanami T., Kano T., Koizumi M. Pathogenic diversity of Citrus tatter leaf virus isolates [J]. Annals of the Plant Pathological Society of Japan,1991,57:74.
[19]Nishio T., Kawai A., Kato M., et al. A sap-transmissible Closterovirus in citrus imported from China and Formosa [J]. Research Bulletin of the Plant Protection Service,1982,18:11-18.
[20]陈集双,张天淼.柑橘碎叶病毒研究[J].微生物学报,1995,35(3)：173-180.
[21]Roistacher C. N., Nauer E. M., Wagner R. L. Transmissibility of Cachexia, Sweet mottle, Psorosis, Tatterleaf and Infectious variegation viruses on knife blades and its prevention [C]. Proceedings of the 8th Conference of the International Organization of Citrus Virologists. Riverside:IOCV,1980:225-229.
[22]Tanner J. D., Kunta M., Da Graca J. V., et al. Evidence of a low rate of seed transmission of Citrus tatter leaf virus in citrus [J]. Phytopathology,2011,101(6): S175-S175.
[23]Martelli G. P., Adams M. J., Kreuze J. F., et al. Family flexiviridae:A case study in virion and genome plasticity [J]. Annual Review of Phytopathology,2007,45: 73-100.
[24]King A. M. Q., Lefkowitz E., Adams M. J., et al. Virus taxonomy:Ninth Report of the International Committee on Taxonomy of Viruses [M]. London:Elsevier, 2011.
[25]Jelkmann W. Cherry virus-A-cDNA cloning of dsRNA, nucleotide-sequence analysis and serology reveal a new plant Capillovirus in sweet cherry [J]. Journal of General Virology,1995,76:2015-2024.
[26]Yoshikawa N., Sasaki E., Kato M., et al. The nucleotide sequence of Apple stem grooving Capillovirus genome [J]. Virology,1992,191(1):98-105.
[27]Komatsu K., Hirata H., Fukagawa T., et al. Infection of Capilloviruses requires subgenomic RNAs whose transcription is controlled by promoter-like sequences conserved among flexiviruses [J]. Virus Research,2012,167(1):8.
[28]Nishio T., Kawai A., Takahashi T., et al. Purification and properties of Citrus tatter leaf virus [J]. Annals of the Phytopathological Society of Japan,1989,55: 254-258.
[29]Zhao L., Hao X., Liu P., et al. Complete sequence of an Apple stem grooving virus (ASGV) isolate from China [J]. Virus Genes,2012,45(3):1-4.
[30]Magome H., Yoshikawa N., Takahashi T. Nucleotide sequence of Citrus tatter leaf virus (isolate li-23) genome [J]. Ann. Phytopathol. Soc. Jpn,1994,60:762-763.
[31]Ohira K., Namba S., Rozanov M., et al. Complete sequence of an infectious full-length cDNA clone of Citrus tatter leaf Capillovirus:Comparative sequence analysis of Capillovirus genomes [J]. Journal of General Virology,1995,76(9): 2305-2309.
[32]Magome H., Yoshikawa N., Takahashi T., et al. Molecular variability of the genomes of Capilloviruses from apple, Japanese pear, European pear, and citrus trees [J]. Phytopathology,1997,87(4):389-396.
[33]Tatineni S., Afunian M. R., Hilf M. E., et al. Molecular characterization of Citrus tatter leaf virus historically associated with Meyer lemon trees:Complete genome sequence and development of biologically active in vitro transcripts [J]. Phytopathology,2009,99(4):423-431.
[34]Bar-Joseph M., Loebenstein G., Cohen J. Partial purification of virus-like particles associated with the Citrus tristeza disease [J]. Phytopathology,1970, 60(1):75-78.
[35]Ohira K., Ito T., Kawai A., et al. Nucleotide sequence of the 3'-terminal region of Citrus tatter leaf virus RNA [J]. Virus Genes,1994,8(2):169-172.
[36]Rochon D. M., Johnston J. C. Infectious transcripts from cloned Cucumber necrosis virus cDNA:Evidence for a bifunctional subgenomic mRNA [J]. Virology,1991,181(2):656-665.
[37]Zhou H., Jackson A. O. Expression of the Barley stripe mosaic virus RNAβ "Triple Gene Block" [J]. Virology,1996,216(2):367-379.
[38]Hirata H., Yamaji Y., Komatsu K., et al. Pseudo-polyprotein translated from the full-length ORF1 of Capillovirus is important for pathogenicity, but a truncated ORF1 protein without variable and CP regions is sufficient for replication [J]. Virus Research,2010,152(1):1-9.
[39]Tatineni S., Afunian M. R., Gowda S., et al. Characterization of the 5'- and 3'-terminal subgenomic RNAs produced by a Capillovirus:Evidence for a CP subgenomic RNA [J]. Virology,2009,385(2):521-528.
[40]Calavan E. C, Christiansen D. W, Roistacher C. N. Symptoms associated with tatter leaf virus infection of Troyer citrange rootstock [J]. Plant Disease,1963, 47:971-975.
[41]Garnsey S. M, Jones J. W. Relationship of symptoms to the presence of tatter-leaf virus in several citrus hosts [C]. Proceedings of the 4th Conference of the International Organization of Citrus Virologists. Riverside:IOCV, 1968:206-212.
[42]何新华,蒋元晖,赵学源等.部分引进柑橘品种的裂皮病和碎叶病鉴定[J].四川果树,1993,21(2)：4-5.
[43]Kawai A., Nishio T. Detection of Citrus tatter leaf virus by enzyme-linked immunosorbent assay (ELISA) [J]. Annals of the Phytopathological Society of Japan,1990,56(3):342-345.
[44]Kawai A., Tsukamoto T., Namba S., et al. Citrus tatter leaf virus:A review of its properties and the development of a serological detection system [J]. Proceedings of the 13th Conference of the International Organization of Citrus Virologists. River-side:IOCV,1996:339-342.
[45]王小凤,黄谊,王文慧等.苹果茎沟病毒的提纯和检测[J].微生物学报,1997,37(4)：258-264.
[46]洪霓,王国平,于济民等.苹果茎沟病毒的分离纯化及血清学检测[J].中国农业科学,1997,30(5)：6-12.
[47]韦晓霞,吴如健.柑橘碎叶病毒的提纯[J].福建果树,1999,108(2)：17-19.
[48]Barba M., Clark M. F. Detection of strains of Apple chlorotic leafspot virus by F(ab)2-based indirect ELISA [C]. XIII International Symposium on Fruit Tree Virus Diseases 193.1985:297-304.
[49]Lovisolo O., Accotto G. P., Masenga V., et al. An isolate of Apple stem grooving virus associated with Cleopatra mandarin fruit intumescence [J]. Fitopatologia Brasileira,2003,28(1):54-58.
[50]黄训才,邓子牛,胡春华等.柑橘主要病毒病的RT-PCR法鉴定[J].湖南农业大学学报,2006,32(3)：273-276.
[51]Hailstones D. L., Bryant K. L., Broadbent P., et al. Detection of Citrus tatter leaf virus with reverse transcription-polymerase chain reaction (RT-PCR) [J]. Australasian Plant Pathology,2000,29(4):240-248.
[52]周常勇,Deborah H., Rachael C等.一种微量、快速抽提柑橘衰退病毒(ctv)核酸应用于RT-PCR扩增的方法[J].福建农业学报,2001,30(增刊)：200.
[53]唐科志,周常勇,王雪峰等.利用RT-PCR检测柑橘碎叶病毒的研究[J].植物病理学报,2005,35(6)：118-119.
[54]Ito T., Ieki H., Ozaki K. Simultaneous detection of six citrus viroids and Apple stem grooving virus from citrus plants by multiplex reverse transcription polymerase chain reaction [J]. Journal of Virological Methods,2002,106(2): 235-239.
[55]Roy A., Fayad A., Barthe G., et al. A multiplex polymerase chain reaction method for reliable, sensitive and simultaneous detection of multiple viruses in citrus trees [J]. Journal of Virological Methods,2005,129(1):47-55.
[56]肖远辉,曾继吾,张秋明等.柑橘衰退病、裂皮病和碎叶病的多重RT-PCR检测方法研究[J].植物病理学报,2007,37(1)：31-35.
[57]Heid C. A., Stevens J., Livak K. J., et al. Real time quantitative PCR [J]. Genome Research,1996,6(10):986-994.
[58]Garrido A., Chapela M. J., Roman B., et al. A new multiplex real-time PCR developed method for Salmonella spp. And listeria monocytogenes detection in food and environmental samples [J]. Food Control,2013,30(1):76-85.
[59]Xiong A. S., Jiang H. H., Zhuang J., et al. Expression and function of a modified AP2/ERF transcription factor from Brassica napus enhances cold tolerance in transgenic Arabidopsis [J]. Molecular Biotechnology,2013,53(2):198-206.
[60]郭立新,向本春,陈红运等.实时荧光RT-PCR一步法检测苹果茎沟病毒[J].植物病理学报,2006,36(1)：57-61.
[61]殷幼平,黄冠军,赵云等.柑橘溃疡病菌实时荧光定量PCR检测与应用[J].植物保护学报,2008,34(6)：607-613.
[62]刘科宏,周常勇,宋震等.运用实时荧光RT-PCR技术检测柑橘碎叶病毒[J].果树学报,2009,26(5)：748-751.
[63]何新华,将元晖.11个柑橘碎叶病毒分离株致病力强弱的比较(初报)[J].云南农业大学学报,1993,8(3)：240-241.
[64]Terauchi H, Magome H., Yoshikawa N., et al. Construction of an infectious cDNA clone of the Apple stem grooving Capillovirus (isolate li-23) genome containing a Cauliflower mosaic virus 35S RNA promoter [J]. Annals of the Phytopathological Society of Japan,1997,63.
[65]孟祥春,向旭,钟云等.砂糖橘上碎叶病毒外壳蛋白基因的克隆及序列分析 [J].果树学报,2007,24(6)：796-798.
[66]Magome H., Yoshikawa N., Takahashi T. Single-strand conformation polymorphism analysis of Apple stem grooving Capillovirus sequence variants [J]. Phytopathology,1999,89(2):136-140.
[67]Gillings M., Broadbent P., Indsto J., et al. Characterisation of isolates and strains of Citrus tristeza closterovirus using restriction analysis of the coat protein gene amplified by the polymerase chain reaction [J]. Journal of Virological Methods, 1993,44(2):305-317.
[68]魏太云,林含新,吴祖建等.应用PCR-RFLP及PCR-sscp技术研究我国水稻条纹病毒RNA4基因间隔区的变异[J].农业生物技术学报,2000,8(1)：41-44.
[69]Navarro L., Civerolo E. L, Juarez J., et al. Improvement therapy methods for citrus germplasm exchange [C]. Proceedings of the 10th Conference of the International Organization of Citrus Virologists. River-side:IOCV,1989:43
[70]何新华,蒋元晖,赵学源等.应用热处理—茎尖嫁接方法培育无碎叶病毒的柑橘[J].果树学报,1993,2：007.
[71]赵小龙,张社南,刘升球.脱除茂谷柑碎叶病毒,天草衰退病毒的试验报告[J].浙江柑橘,2006,23(3)：19-20.
[72]何新华,赵学源.柑橘碎叶病毒对八种不同来源枳砧锦橙的反应及脱毒研究[J].广西植保,1996,9(2)：4-6.
[73]Boyer J. C., Haenni A. L. Infectious transcripts and cDNA clones of RNA viruses [J]. Virology,1994,198(2):415-426.
[74]Taniguchi T., Palmieri M., Weissman C. Qβ DNA-containing hybrid plasmids giving rise to Qβ phage formation in the bacterial host [J]. Nature,1978,274 223-228.
[75]Ahlquist P., French R., Janda M., et al. Multicomponent RNA plant virus infection derived from cloned viral cDNA [J]. Proceedings of the National Academy of Sciences,1984,81(22):7066-7070.
[76]Petty I. T, Hunter B. G, Wei N., et al. Infectious Barley stripe mosaic virus RNA transcribed in vitro from full-length genomic cDNA clones [J]. Virology,1989, 171(2):342-349.
[77]Hayes R. J, Buck K. W. Infectious Cucumber mosaic virus RNA transcribed in vitro from clones obtained from cDNA amplified using the polymerase chain reaction [J]. Journal of General Virology,1990,71(11):2503-2508.
[78]Timpe U., Kuhne T. In vitro transcripts of a full-length cDNA of a naturally deleted RNA2 of Barley mild mosaic virus (BAMMV) replicate in BAMMV-infected plants [J]. Journal of General Virology,1995,76(10): 2619-2623.
[79]Meyer M., Dessens J. T.35 S promoter-driven cDNAs of Barley mild mosaic virus RNA1 and RNA2 are infectious on barley plants [J]. Journal of General Virology, 1997,78(12):3147-3151.
[80]Riechmann J. L, Lain S., Garcia JA. Infectious in vitro transcripts from a Plum poxpotyvirus cDNA clone [J]. Virology,1990,177(2):710.
[81]Sanchez F., Martnez-Herrera D., Aguilar I., et al. Infectivity of Turnip mosaic potyvirus cDNA clones and transcripts on the systemic host Arabidopsis thaliana and local lesion hosts [J]. Virus Research,1998,55(2):207-219.
[82]Kim K. S., Oh H. Y., Suranto S., et al. Infectivity of in vitro transcripts of Johnsongrass mosaic potyvirus full-length cDNA clones in maize and sorghum [J]. Archives of Virology,2003,148(3):563-574.
[83]薛朝阳,周雪平,陈青等.一种病毒侵染性全长cDNA克隆的快速构建方法[J].生物化学与生物物理学报,2000,32(3)：270-274.
[84]Dagless E. M, Shintaku M. H, Nelson R. S, et al. A CaMV 35S promoter driven cDNA clone of Tobacco mosaic virus can infect host plant tissue despite being uninfectious when manually inoculated onto leaves [J]. Archives of Virology, 1997,142(1):183-191.
[85]Yang S. J, Revers F., Souche S., et al. Construction of full-length cDNA clones of Lettuce mosaic virus (LMV) and the effects of intron-insertion on their viability in Escherichia coli and on their infectivity to plants [J]. Archives of Virology, 1998,143(12):2443-2451.
[86]Saldarelli P., Dell'Orco M., Minafra A. Infectious cDNA clones of two grapevine viruses [J]. Archives of Virology,2000,145(2):397-405.
[87]Rice C. M, Grakoui A., Galler R., et al. Transcription of infectious yellow fever RNA from full-length cDNA templates produced by in vitro ligation [J]. The New Biologist,1989,1(3):285.
[88]Forns X., Bukh J., Purcell R. H., et al. How Escherichia coli can bias the results of molecular cloning:Preferential selection of defective genomes of Hepatitis C virus during the cloning procedure [J]. Proceedings of the National Academy of Sciences,1997,94(25):13909-13914.
[89]Fakhfakh H., Vilaine F., Makni M., et al. Cell-free cloning and biolistic inoculation of an infectious cDNA of Potato virus Y [J]. Journal of General Virology,1996,77(3):519-523.
[90]Jakab G., Droz E., Brigneti G., et al. Infectious in vivo and in vitro transcripts from a full-length cDNA clone of PVY-n605, a Swiss necrotic isolate of potato virus y [J]. Journal of General Virology,1997,78(12):3141-3145.
[91]Sriburi R., Keelapang P., Duangchinda T., et al. Construction of infectious Dengue 2 virus cDNA clones using high copy number plasmid [J]. Journal of Virological Methods,2001,92(1):71-82.
[92]Skotnicki M. L, Ding S.W., Mackenzie A. M, et al. Infectious Eggplant mosaic tymovirus and Ononis yellow mosaic tymovirus from cloned cDNA [J]. Archives of Virology,1993,131(1):47-60.
[93]Sumiyoshi H., Hoke C. H., Trent D. W. Infectious Japanese encephalitis virus RNA can be synthesized from in vztro-ligated cDNA templates [J]. Journal of Virology,1992,66(9):5425-5431.
[94]Johansen I. E. Intron insertion facilitates amplification of cloned virus cDNA in Escherichia coli while biological activity is reestablished after transcription in vivo [J]. Proceedings of the National Academy of Sciences,1996,93(22): 12400-12405.
[95]Olsen B. S., Johansen I. E. Nucleotide sequence and infectious cDNA clone of the 11 isolates of Pea seed-borne mosaic potyvirus [J]. Archives of Virology, 2001,146(1):15-25.
[96]Maiss E., Timpe U., Brisske-Rode A., et al. Infectious in vivo transcripts of a Plum pox potyvirus full-length cDNA clone containing the Cauliflower mosaic virus 35S RNA promoter [J]. Journal of General Virology,1992,73(3):709-713.
[97]Vancanneyt G., Schmidt R., O'Connor-Sanchez A., et al. Construction of an intron-containing marker gene:Splicing of the intron in transgenic plants and its use in monitoring early events in Agrobacterium-mediated plant transformation [J]. Molecular and General Genetics MGG,1990,220(2):245-250.
[98]Gonzalez J. M., Penzes Z., Almazan F., et al. Stabilization of a full-length infectious cDNA clone of transmissible Gastroenteritis coronavirus by insertion of an intron [J]. Journal of Virology,2002,76(9):4655-4661.
[99]Yamshchikov V., Mishin V., Cominelli F. A new strategy in design of (+) RNA virus infectious clones enabling their stable propagation in E. Coli [J]. Virology, 2001,281(2):272-280.
[100]Polo S., Ketner G., Levis R., et al. Infectious RNA transcripts from full-length Dengue virus type 2 cDNA clones made in yeast [J]. Journal of Virology,1997, 71(7):5366-5374.
[101]MacFarlane S. A., Wallis C. V., Taylor S. C., et al. Construction and analysis of infectious transcripts synthesized from full-length cDNA clones of both genomic RNAs of Pea early browning virus [J]. Virology,1991,182(1):124-129.
[102]Flasinski S., Cassidy B. G. Potyvirus aphid transmission requires helper component and homologous coat protein for maximal efficiency [J]. Archives of Virology,1998,143(11):2159-2172.
[103]Zhang F., Huang Q., Ma W., et al. Amplification and cloning of the full-length genome of Japanese encephalitis virus by a novel long RT-PCR protocol in a cosmid vector [J]. Journal of Virological Methods,2001,96(2):171-182.
[104]Joshi A., Jeang KT. Reduction in growth temperature minimizes instability of large plasmids containing HIV-1 proviral genomes [J]. Biotechniques,1993, 14(6):880-884.
[105]Domingo E. Mutation rates and rapid evolution of RNA viruses [J]. The Evolutionary Biology of Viruses.,1994:161-184.
[106]Shimotohno K., Kodama Y., Hashimoto J., et al. Importance of 5'-terminal blocking structure to stabilize mRNA in eukaryotic protein synthesis [J]. Proceedings of the National Academy of Sciences,1977,74(7):2734-2738.
[107]Dolja V. V, McBride H. J., Carrington J.C. Tagging of plant Potyvirus replication and movement by insertion of beta-glucuronidase into the viral polyprotein [J]. Proceedings of the National Academy of Sciences,1992,89(21):10208-10212.
[108]Gal-On A., Antignus Y, Rosner A., et al. Infectious in vitro RNA transcripts derived from cloned cDNA of the Cucurbit potyvirus, Zucchini yellow mosaic virus [J]. Journal of General Virology,1991,72(11):2639-2643.
[109]Sivakumaran K., Fowler B. C., Hacker D.L. Identification of viral genes required for cell-to-cell movement of Southern bean mosaic virus [J]. Virology,1998, 252(2):376-386.
[110]Angenent G. C, Posthumus E., Bol J. F. Biological activity of transcripts synthesized in vitro from full-length and mutated DNA copies of Tobacco rattle virus RNA2 [J]. Virology,1989,173(1):68-76.
[111]Hamilton W. D. O., Baulcombe D. C. Infectious RNA produced by in vitro transcription of a full-length Tobacco rattle virus RNA-1 cDNA [J]. Journal of General Virology,1989,70(4):963-968.
[112]Huang M., Koh D. C. Y., Weng L. J., et al. Complete nucleotide sequence and genome organization of Hibiscus chlorotic ringspot virus, a new member of the genus Carmovirus:Evidence for the presence and expression of two novel open reading frames [J]. Journal of Virology,2000,74(7):3149-3155.
[113]Mayo M. A., Barker H., Robinson D. J., et al. Evidence that Potato leafroll virus RNA is positive-stranded, is linked to a small protein and does not contain polyadenylate [J]. Journal of General Virology,1982,59(1):163-167.
[114]Tobin G. J., Young D. C., Flanegan J. B. Self-catalyzed linkage of Poliovirus terminal protein VPG to Poliovirus RNA [J]. Cell,1989,59(3):511-519.
[115]Sit T. L., AbouHaidar M. G. Infectious RNA transcripts derived from cloned cDNA of Papaya mosaic virus:Effect of mutations to the capsid and polymerase proteins [J]. Journal of General Virology,1993,74(6):1133-1140.
[116]Jackson R. J., Standart N. Do the poly (A) tail and 3'untranslated region control mRNA translation? [J]. Cell,1990,62(1):15-24.
[117]Tsai C. H., Cheng C. P., Peng C. W., et al. Sufficient length of a poly (A) tail for the formation of a potential pseudoknot is required for efficient replication of Bamboo mosaic potexvirus RNA [J]. Journal of Virology,1999,73(4): 2703-2709.
[118]Chiang C. H., Yeh S. D. Infectivity assays of in vitro and in vivo transcripts of Papaya ringspot potyvirus [J]. Botanical Bulletin of Academia Sinica,1997,38: 153-163.
[119]Barnes W. M. PCR amplification of up to 35-kb DNA with high fidelity and high yield from Lambda bacteriophage templates [J]. Proceedings of the National Academy of Sciences,1994,91(6):2216-2220.
[120]Heaton L. A., Carrington J. C., Morris T. J. Turnip crinkle virus infection from RNA synthesized in vitro [J]. Virology,1989,170(1):214-218.
[121]Bujarski Jozef J., Miller W. A. Use of in vitro transcription to study gene expression and replication of spherical, positive sense RNA plant viruses [J]. Genetic Engineering with Plant Viruses,1992:115.
[122]Commandeur U., Jarausch W., Li Y., et al. cDNAs of Beet necrotic yellow vein virus RNAs 3 and 4 are rendered biologically active in a plasmid containing the Cauliflower mosaic virus 35S promoter [J]. Virology,1991,185(1):493-495.
[123]Mori M., Mise K., Kobayashi K., et al. Infectivity of plasmids containing Brome mosaic virus cDNA linked to the Cauliflower mosaic virus 35S RNA promoter [J]. Journal of General Virology,1991,72(2):243-246.
[124]Neeleman L., Van Der Vossen E. A. G., Bol J. F. Infection of tobacco with Alfalfa mosaic virus cDNAs sheds light on the early function of the coat protein [J]. Virology,1993,196(2):883-887.
[125]Zaitlin M., Palukaitis P. Advances in understanding plant viruses and virus diseases [J]. Annual Review of Phytopathology,2000,38(1):117-143.
[126]Lopez-Moya J. J., Garcia J. A. Construction of a stable and highly infectious intron-containing cDNA clone of Plum pox potyvirus and its use to infect plants by particle bombardment [J]. Virus Research,2000,68(2):99-107.
[127]Ding S. W., Rathjen J. P, Li W. X., et al. Efficient infection from cDNA clones of Cucumber mosaic cucumovirus RNAs in a new plasmid vector [J]. Journal of General Virology,1995,76(2):459-464.
[128]Weber H., Haeckel P., Pfitzner A. J. A cDNA clone of Tomato mosaic virus is infectious in plants [J]. Journal of Virology,1992,66(6):3909-3912.
[129]Gal-On A., Meiri E., Huet H., et al. Particle bombardment drastically increases the infectivity of cloned DNA of Zucchini yellow mosaic potyvirus [J]. Journal of General Virology,1995,76(12):3223-3227.
[130]Yamashita T., Iida A., Morikawa H. Evidence that more than 90% of β-glucuronidase-expressing cells after particle bombardment directly receive the foreign gene in their nucleus [J]. Plant Physiology,1991,97(2):829-831.
[131]Grimsley N., Hohn B., Hohn T., et al. "Agroinfection," an alternative route for viral infection of plants by using the Ti plasmid [J]. Proceedings of the National Academy of Sciences,1986,83(10):3282-3286.
[132]Lindbo J. A. High-efficiency protein expression in plants from agroinfection-compatible Tobacco mosaic virus expression vectors [J]. BMC Biotechnology, 2007,7(1):1-11.
[133]Ratcliff F., Martin-Hernandez A. M., Baulcombe D. C. Tobacco rattle virus as a vector for analysis of gene function by silencing [J]. Plant Journal,2001,25(2): 237-245.
[134]Shi B. J., Ding S. W., Symons R. H. Plasmid vector for cloning infectious cDNAs from plant RNA viruses:High infectivity of cDNA clones of Tomato aspermy cucumovirus [J]. Journal of General Virology,1997,78(5):1181-1185.
[135]Leiser R. M., Ziegler-Graff V., Reutenauer A., et al. Agroinfection as an alternative to insects for infecting plants with Beet western yellows luteovirus [J]. Proceedings of the National Academy of Sciences,1992,89(19):9136-9140.
[136]Grimsley N. H., Ramos C., Hein T.; et al. Meristematic tissues of maize plants are most susceptible to agroinfection with Maize streak virus [J]. Nature Biotechnology,1988,6(2):185-189.
[137]Matthew L. RNA i for plant functional genomics [J]. Comparative and Functional Genomics,2004,5(3):240-244.
[138]Van Kammen A. Virus-induced gene silencing in infected and transgenic plants [J]. Trends in Plant Science,1997,2(11):409-411.
[139]Kumagai M. H., Donson J., Della-Cioppa G., et al. Cytoplasmic inhibition of carotenoid biosynthesis with virus-derived RNA [J]. Proceedings of the National Academy of Sciences,1995,92(5):1679-1683.
[140]Godge M. R., Purkayastha A., Dasgupta I., et al. Virus-induced gene silencing for functional analysis of selected genes [J]. Plant Cell Reports,2008,27(2): 209-219.
[141]Burch-Smith T. M., Anderson J. C., Martin G. B., et al. Applications and advantages of virus-induced gene silencing for gene function studies in plants [J]. Plant Journal,2004,39(5):734-746.
[142]Baulcombe D. C. Fast forward genetics based on virus-induced gene silencing [J]. Current Opinion in Plant Biology,1999,2(2):109-113.
[143]Becker A., Lange M. Vigs-genomics goes functional [J]. Trends in Plant Science, 2010,15(1):1-4.
[144]Purkayastha A., Dasgupta I. Virus-induced gene silencing:A versatile tool for discovery of gene functions in plants [J]. Plant Physiology and Biochemistry, 2009,47(11):967-976.
[145]Senthil-Kumar M., Mysore K. S. New dimensions for VIGS in plant functional genomics [J]. Trends in Plant Science,2011,16(12):656-665.
[146]Baulcombe D. RNA silencing in plants [J]. Nature,2004,431(7006):356-363.
[147]Meyer P., Heidmann I., Niedenhof I. Differences in DNA-methylation are associated with a paramutation phenomenon in transgenic petunia [J]. The Plant Journal,2002,4(1):89-100.
[148]Llave C. Virus-derived small interfering RNAs at the core of plant-virus interactions [J]. Trends in Plant Science,2010,15(12):701-707.
[149]Donaire L., Barajas D., Martinez-Garcia B., et al. Structural and genetic requirements for the biogenesis of Tobacco rattle virus-derived small interfering RNAs [J]. Journal of Virology,2008,82(11):5167-5177.
[150]Qu F., Ye X., Morris T. J. Arabidopsis DRB4,AGO1,AGO7, and RDR6 participate in a DCL4-initiated antiviral RNA silencing pathway negatively regulated by DCL1 [J]. Proceedings of the National Academy of Sciences,2008,105(38): 14732-14737.
[151]Brodersen P., Sakvarelidze-Achard L., Bruun-Rasmussen M., et al. Widespread translational inhibition by plant miRNAs and siRNAs [J]. Science,2008, 320(5880):1185-1190.
[152]Zheng X., Zhu J., Kapoor A., et al. Role of ArabidopsisAGO6 in siRNA accumulation, DNA methylation and transcriptional gene silencing [J]. The EMBO journal,2007,26(6):1691-1701.
[153]Sasaki S., Yamagishi N., Yoshikawa N. Efficient virus-induced gene silencing in apple, pear and Japanese pear using Apple latent spherical virus vectors [J]. Plant Methods,2011,7(1):15.
[154]van der Linde K., Kastner C., Kumlehn J., et al. Systemic virus-induced gene silencing allows functional characterization of maize genes during biotrophic interaction with ustilago maydis [J]. New Phytologist,2011,189(2):471-483.
[155]Zhang C., Bradshaw J. D., Whitham S. A., et al. The development of an efficient multipurpose Bean pod mottle virus viral vector set for foreign gene expression and RNA silencing [J]. Plant Physiology,2010,153(1):52-65.
[156]Aguero J., Ruiz-Ruiz S., Vives M. D., et al. Development of viral vectors based on Citrus leaf blotch virus to express foreign proteins or analyze gene function in citrus plants [J]. Molecular Plant-Microbe Interact,2012,25(10):1326-1337.
[157]Nagamatsu A., Masuta C., Senda M., et al. Functional analysis of soybean genes involved in flavonoid biosynthesis by virus-induced gene silencing [J]. Plant Biotechnology Journal,2007,5(6):778-790.
[158]Lu H. C., Chen H. H., Tsai W. C., et al. Strategies for functional validation of genes involved in reproductive stages of orchids [J]. Plant Physiology,2007, 143(2):558-569.
[159]Kurth E. G., Peremyslov V. V., Prokhnevsky A. I., et al. Virus-derived gene expression and RNA interference vector for grapevine [J]. Journal of Virology, 2012,86(11):6002-6009.
[160]Faivre-Rampant O., Gilroy E. M., Hrubikova K., et al. Potato virus X-induced gene silencing in leaves and tubers of potato [J]. Plant Physiology,2004,134(4): 1308-1316.
[161]Gammelgard E., Mohan M., Valkonen J. P. T. Potyvirus-induced gene silencing: The dynamic process of systemic silencing and silencing suppression [J]. Journal of General Virology,2007,88(8):2337-2346.
[162]Constantin G. D., Gr(?)nlund M., Johansen I. E., et al. Virus-induced gene silencing (VIGS) as a reverse genetic tool to study development of symbiotic root nodules [J]. Molecular Plant-Microbe Interactions,2008,21(6):720-727.
[163]Naylor M., Reeves J., Cooper J. I., et al. Construction and properties of a gene-silencing vector based on Poplar mosaic virus (genus carlavirus) [J]. Journal of Virological Methods,2005,124(1-2):27-36.
[164]Vaistij F. E., Jones L. Compromised virus-induced gene silencing in RDR6-deficient plants [J]. Plant Physiology,2009,149(3):1399-1407.
[165]Varallyay E., Lichner Z., Safrany J., et al. Development of a virus induced gene silencing vector from a legumes infecting tobamovirus [J]. Acta Biologica Hungarica,2010,61(4):457-469.
[166]Hiriart J. B., Aro E. M., Lehto K. Dynamics of the VIGS-mediated chimeric silencing of the Nicotiana benthamiana chlh gene and of the Tobacco mosaic virus vector [J]. Molecular Plant-Microbe Interactions,2003,16(2):99-106.
[167]Gao Y., Zhang Y. L., Zhang X. F., et al. Development and optimization of Tobacco necrosis virus a induced gene silencing in Nicotiana benthamiana [J]. Progress in Biochemistry and Biophysics,2011,38(10):919-928.
[168]Pignatta D., Kumar P., Turina M., et al. Quantitative analysis of efficient endogenous gene silencing in Nicotiana benthamiana plants using Tomato bushy stunt virus vectors that retain the capsid protein gene [J]. Molecular Plant-Microbe Interactions,2007,20(6):609-618.
[169]Burch-Smith T. M., Schiff M., Liu Y., et al. Efficient virus-induced gene silencing in Arabidopsis [J]. Plant Physiology,2006,142(1):21-27.
[170]Fu D. Q., Zhu B. Z., Zhu H. L., et al. Enhancement of virus-induced gene silencing in tomato by low temperature and low humidity [J]. Molecules and Cells,2006,21(1):153.
[171]Ryu C. M., Anand A., Kang L., et al. Agrodrench:A novel and effective agroinoculation method for virus-induced gene silencing in roots and diverse solanaceous species [J]. The Plant Journal,2004,40(2):322-331.
[172]Fu D. Q., Zhu B. Z., Zhu H. L., et al. Virus-induced gene silencing in tomato fruit [J]. The Plant Journal,2005,43(2):299-308.
[173]Pflieger S., Blanchet S., Camborde L., et al. Efficient virus-induced gene silencing in Arabidopsis using a 'one-step'TYMV-derived vector [J]. The Plant Journal,2008,56(4):678-690.
[174]Krenz B., Wege C., Jeske H. Cell-free construction of disarmed Abutilon mosaic virus-based gene silencing vectors [J]. Journal of Virological Methods,2010, 169(1):129-137.
[175]Fofana I. B. F., Sangare A., Collier R., et al. A geminivirus-induced gene silencing system for gene function validation in cassava [J]. Plant Molecular Biology,2004,56(4):613-624.
[176]Golenberg E. M., Sather D. N., Hancock Leandria C., et al. Development of a gene silencing DNA vector derived from a broad host range Geminivirus [J]. Plant Methods,2009,5(1):1-14.
[177]Turnage M. A., Muangsan N., Peele C. G., et al. Geminivirus-based vectors for gene silencing in Arabidopsis [J]. The Plant Journal,2002,30(1):107-114.
[178]Tuttle J. R., Idris A. M, Brown J. K., et al. Geminivirus-mediated gene silencing from Cotton leaf crumple virus is enhanced by low temperature in cotton [J]. Plant Physiology,2008,148(1):41-50.
[179]Muruganantham M., Moskovitz Y., Haviv S., et al. Grapevine virusa-mediated gene silencing in Nicotiana benthamiana and Vitis vinifera [J]. Journal of Virological Methods,2009,155(2):167-74.
[180]del Rosario A. J. M., del Carmen R. G. M., Lopez M. G., et al. Virus-induced silencing of comt, pamt and kas genes results in a reduction of capsaicinoid accumulation in chili pepper fruits [J]. Planta,2008,227(3):681-695.
[181]Pandey P., Choudhury N. R., Mukherjee S. K. A geminiviral amplicon (VA) derived from Tomato leaf curl virus (ToLCV) can replicate in a wide variety of plant species and also acts as a VIGS vector [J]. Virology Journal,2009,6(1): 1-13.
[182]Gossele V., Fache I., Meulewaeter F., et al. SVISS-a novel transient gene silencing system for gene function discovery and validation in tobacco plants [J]. The Plant Journal,2002,32(5):859-866.
[183]Cai X., Wang C., Xu Y., et al. Efficient gene silencing induction in tomato by a viral satellite DNA vector [J]. Virus Research,2007,125(2):169-175.
[184]Tao X., Zhou X. A modified viral satellite DNA that suppresses gene expression in plants [J]. Plant Journal,2004,38(5):850-860.
[185]Qian Y. J., Mugiira R. B., Zhou X. P. A modified viral satellite DNA-based gene silencing vector is effective in association with Heterologous begomoviruses [J]. Virus Research,2006,118(1):136-142.
[186]Huang C. J., Xie Y., Zhou X. P. Efficient virus-induced gene silencing in plants using a modified Geminivirus DNA1 component [J]. Plant Biotechnology Journal, 2009,7(3):254-265.
[187]Ruiz M. T., Voinnet O., Baulcombe D. C. Initiation and maintenance of virus-induced gene silencing [J]. The Plant Cell Online,1998,10(6):937-946.
[188]Kjemtrup S., Sampson K. S., Peele C. G., et al. Gene silencing from plant DNA carried by a Geminivirus [J]. The Plant Journal,2002,14(1):91-100.
[189]Zhou X. P., Huang C. J. Virus-induced gene silencing using Begomovirus satellite molecules [J]. Methods Mol Biol,2012,894:57-67.
[190]Holzberg S., Brosio P., Gross C., et al. Barley stripe mosaic virus-induced gene silencing in a monocot plant [J]. Plant Journal,2002,30(3):315-327.
[191]Scofield S. R., Brandt A. S. Virus-induced gene silencing in hexaploid wheat using Barley stripe mosaic virus vectors [J]. Methods in molecular biology,2012, 894:93-112.
[192]Scofield S. R., Huang L., Brandt A. S., et al. Development of a virus-induced gene-silencing system for hexaploid wheat and its use in functional analysis of the lr21-mediated leaf rust resistance pathway [J]. Plant Physiology,2005, 138(4):2165-2173.
[193]Constantin G. D., Krath B. N., MacFarlane S. A., et al. Virus-induced gene silencing as a tool for functional genomics in a legume species [J]. Plant Journal, 2004,40(4):622-631.
[194]Zhang C., Ghabrial S. A. Development of Bean pod mottle virus-based vectors for stable protein expression and sequence-specific virus-induced gene silencing in soybean [J]. Virology,2006,344(2):401-11.
[195]Zhang C., Yang C., Whitham S. A., et al. Development and use of an efficient DNA-based viral gene silencing vector for soybean [J]. Molecular Plant-Microbe Interactions,2009,22(2):123-131.
[196]Ding X. S., Schneider W. L., Chaluvadi S. R., et al. Characterization of a brome mosaic virus strain and its use as a vector for gene silencing in monocotyledonous hosts [J]. Molecular Plant-Microbe Interact,2006,19(11): 1229-1239.
[197]Idris A. M., Ktenz B., Tuttle J. R., et al. Construction of a virus-induced gene silencing (VIGS)vector for cotton using Cotton leaf crumple virus and a fragment of the cotton phytoene desaturase [J]. Phytopathology,2008,98(6):S70-S70.
[198]Jones A. L., Thomas C. L., Maule A. J. De novo methylation and co-suppression induced by a cytoplasmically replicating plant RNA virus [J]. The EMBO Journal, 1998,17(21):6385-6393.
[199]Kanazawa A., Inaba J., Shimura H., et al. Virus-mediated efficient induction of epigenetic modifications of endogenous genes with phenotypic changes in plants [J]. The Plant Journal,2011,65(1):156-168.
[200]Sonoda S., Nishiguchi M. Delayed activation of post-transcriptional gene silencing and de novo transgene methylation in plants with the coat protein gene of Sweet potato feathery mottle potyvirus [J]. Plant Science,2000,156(2): 137-144.
[201]Vainstein A., Marton I., Zuker A., et al. Permanent genome modifications in plant cells by transient viral vectors [J]. Trends in Biotechnology,2011,29(8): 363-369.
[202]Kim S., Kim J. S. Targeted genome engineering via zinc finger nucleases [J]. Plant Biotechnology Reports,2011,5(1):9-17.
[203]Meng Y., Moscou M. J., Wise R. P. Blufensinl negatively impacts basal defense in response to Barley powdery mildew [J]. Plant Physiology,2009,149(1): 271-285.
[204]Liu Y., Schiff M., Dinesh-Kumar S. P. Virus-induced gene silencing in tomato [J]. Plant Journal,2002,31(6):777-786.
[205]Dong Y., Burch-Smith T. M., Liu Y., et al. A ligation-independent cloning Tobacco rattle virus vector for high-throughput virus-induced gene silencing identifies roles for NbMADS4-1 and-2 in floral development [J]. Plant Physiology, 2007,145(4):1161-1170.
[206]Tang Y, Wang F., Zhao J., et al. Virus-based microRNA expression for gene functional analysis in plants [J]. Plant Physiology,2010,153(2):632-641.
[207]Lacomme C. Milestones in the development and applications of plant virus vector as gene silencing platforms [J]. Current Topics in Microbiology and Immunology, 2011,10.1007/82_2011_186.
[208]Valentine T. A., Randall E., Wypijewski K., et al. Delivery of macromolecules to plant parasitic nematodes using a Tobacco rattle virus vector [J]. Plant Biotechnology Journal,2007,5(6):827-834.
[209]Dubreuil G., Magliano M., Dubrana M. P., et al. Tobacco rattle virus mediates gene silencing in a plant parasitic root-knot nematode [J]. Journal of Experimental Botany,2009,60(14):4041-4050.
[210]Hein I., Barciszewska-Pacak M., Hrubikova K., et al. Virus-induced gene silencing-based functional characterization of genes associated with powdery mildew resistance in barley [J]. Plant Physiology,2005,138(4):2155-2164.
[211]Goodin M. M., Zaitlin D., Naidu R. A., et al. Nicotiana benthamiana:Its history and future as a model for plant-pathogen interactions [J]. Molecular Plant-Microbe Interactions,2008,21(8):1015-1026.
[212]Yang S. J., Carter S. A., Cole A. B., et al. A natural variant of a host RNA-dependent RNA polymerase is associated with increased susceptibility to viruses by Nicotiana benthamiana [J]. Proceedings of the National Academy of Sciences of the United States of America,2004,101(16):6297-6302.
[213]Thomas C. L., Jones L., Baulcombe D. C., et al. Size constraints for targeting post-transcriptional gene silencing and for RNA-directed methylation in Nicotiana benthamiana using a Potato virus X vector [J]. The Plant Journal,2001, 25(4):417-425.
[214]Xu P., Zhang Y., Kang L., et al. Computational estimation and experimental verification of off-target silencing during posttranscriptional gene silencing in plants [J]. Plant Physiology,2006,142(2):429-440.
[215]Senthil-Kumar M., Mysore K. S. Caveat of RNAi in plants:The off-target effect [J]. RNAi and Plant Gene Function Analysis,2011:13-25.
[216]Senthil-Kumar M., Hema R., Anand A., et al. A systematic study to determine the extent of gene silencing in Nicotiana benthamiana and other Solanaceae Species when heterologous gene sequences are used for virus-induced gene silencing [J]. New Phytologist,2007,176(4):782-791.
[217]Jacob S. S., Vanitharani R., Karthikeyan A. S., et al. Mungbean yellow mosaic virus-Vi agroinfection by codelivery of DNA A and DNA B from one Agrobacterium strain [J]. Plant disease,2003,87(3):247-251.
[218]Liu E., Page J. E. Optimized cDNA libraries for virus-induced gene silencing (VIGS) using Tobacco rattle virus [J]. Plant Methods,2008,4(5):1-13.
[219]Rodrigo G., Carrera J., Jaramillo A., et al. Optimal viral strategies for bypassing RNA silencing [J]. Journal of The Royal Society Interface,2011,8(55):257-268.
[220]Lacomme C, Hrubikova K. Enhancement of virus-induced gene silencing through viral-based production of inverted-repeats [J]. The Plant Journal, 2003,34(4):543-553.
[221]Molnar A., Csorba T., Lakatos L., et al. Plant virus-derived small interfering RNAs originate predominantly from highly structured single-stranded viral RNAs [J]. Journal of Virology,2005,79(12):7812-7818.
[222]Szittya G., Silhavy D., Molnar A., et al. Low temperature inhibits RNA silencing-mediated defence by the control of siRNA generation [J]. The EMBO Journal,2003,22(3):633-640.
[223]Orzaez D., Mirabel S., Wieland W. H., et al. Agroinjection of tomato fruits. A tool for rapid functional analysis of transgenes directly in fruit [J]. Plant Physiology,2006,140(1):3-11.
[224]Romero I., Tikunov Y., Bovy A. Virus-induced gene silencing in detached tomatoes and biochemical effects of phytoene desaturase gene silencing [J]. Journal of Plant Physiology,2011,168(10):1129-1135.
[225]Unver T., Budak H. Virus-induced gene silencing, a post transcriptional gene silencing method [J]. International Journal of Plant Genomics,2009:1986-1980.
[226]Lucioli A., Noris E., Brunetti A., et al. Tomato yellow leaf curl Sardinia virus rep-derived resistance to homologous and heterologous Geminiviruses occurs by different mechanisms and is overcome if virus-mediated transgene silencing is activated [J]. Journal of Virology,2003,77(12):6785-6798.
[227]Lu R., Malcuit I., Moffett P., et al. High throughput virus-induced gene silencing implicates heat shock protein 90 in plant disease resistance [J]. The EMBO Journal,2003,22(21):5690-5699.
[228]Chen J. C., Jiang C. Z., Gookin T. E., et al. Chalcone synthase as a reporter in virus-induced gene silencing studies of flower senescence [J]. Plant Molecular Biology,2004,55(4):521-530.
[229]Huang C.J., Qian Y.J., Li Z.H., et al. Virus-induced gene silencing and its application in plant functional genomics [J]. Science China Life Sciences,2012, 55(2):99-108.
[230]Liu Y., Schiff M., Czymmek K., et al. Autophagy regulates programmed cell death during the plant innate immune response [J]. Cell,2005,121(4):567-577'.
[231]Caplan J.L., Mamillapalli P., Burch-Smith T.M., et al. Chloroplastic protein NRIP1 mediates innate immune receptor recognition of a viral effector [J]. Cell, 2008,132(3):449-462.
[232]Gronlund M., Olsen A., Johansen E. I., et al. Protocol:Using virus-induced gene silencing to study the arbuscular mycorrhizal symbiosis in Pisum sativum [J]. Plant Methods,2010,6(1):28.
[233]Zhu X., Dinesh-Kumar S. P. Virus-induced gene silencing as a tool to identify host genes affecting viral pathogenicity [J]. Plant Virology Protocols,2008,451: 641-648.
[234]Cloutier S., McCallum B. D., Loutre C., et al. Leaf rust resistance gene IR1, isolated from bread wheat (Triticum aestivum 1.) is a member of the large psr567 gene family [J]. Plant Molecular Biology,2007,65(1):93-106.
[235]Borras-Hidalgo O., Thomma B. P. H. J., Collazo C., et al. EIL2 transcription factor and glutathione synthetase are required for defense of tobacco against Tobacco blue mold [J]. Molecular Plant-Microbe Interactions,2006,19(4): 399-406.
[236]Ekengren S.K., Liu Y., Schiff M., et al. Two MAPK cascades, NPR1, and TGA transcription factors play a role in Pto-mediated disease resistance in tomato [J]. The Plant Journal,2003,36(6):905-917.
[237]Sharma P. C., Ito A., Shimizu T., et al. Virus-induced silencing of WIPK and SIPK genes reduces resistance to a bacterial pathogen, but has no effect on the INF1-induced hypersensitive response (HR) in Nicotiana benthamiana [J]. Molecular Genetics and Genomics,2003,269(5):583-591.
[238]Maimbo M., Ohnishi K., Hikichi Y., et al. S-glycoprotein-like protein regulates defense responses in Nicotiana plants against Ralstonia solanacearum [J]. Plant Physiology,2010,152(4):2023-2035.
[239]Lee D. H., Choi H. W., Hwang B. K. The pepper E3 ubiquitin ligase RING1 gene, CaRINGl, is required for cell death and the salicylic acid-dependent defense response [J]. Plant Physiology,2011,156(4).
[240]Melech-Bonfil S., Sessa G. The SLMKK2 and SLMPK2 genes play a role in tomato disease resistance to Xanthomonas campestris pv. Vesicatoria [J]. Plant Signaling & Behavior,2011,6(1):154-156.
[241]Mantelin S., Peng H. C., Li B., et al. The receptor-like kinase SLSERK1 is required for Mi-1-mediated resistance to potato aphids in tomato [J]. The Plant Journal,2011,67(3):459-471.
[242]Lou Y., Baldwin I. T. Silencing of a germin-like gene in Nicotiana attenuata improves performance of native herbivores [J]. Plant Physiology,2006,140(3): 1126-1136.
[243]Heinrich M., Baldwin I. T., Wu J. Two mitogen-activated protein kinase kinases, MKK1 and MEK2, are involved in wounding-and specialist Lepidopteran herbivore manduca sexta-induced responses in nicotiana attenuata [J]. Journal of Experimental Botany,2011,62(12):4355-4365.
[244]Guo Y., Huang C., Xie Y., et al. A tomato glutaredoxin gene SLGX1 regulates plant responses to oxidative, drought and salt stresses [J]. Planta,2010,232(6): 1499-1509.
[245]Re D. A., Dezar C. A., Chan R. L., et al. Nicotiana attenuata NaHD20 plays a role in leaf ABA accumulation during water stress, benzylacetone emission from flowers, and the timing of bolting and flower transitions [J]. Journal of Experimental Botany,2011,62(1):155-166.
[246]Lee S. C., Choi D. S., Hwang I. S., et al. The pepper oxidoreductase CAOXR1 interacts with the transcription factor caravl and is required for salt and osmotic stress tolerance [J]. Plant Molecular Biology,2010,73(4):409-424.
[247]Senthil-Kumar M., Govind G., Kang L., et al. Functional characterization of Nicotiana benthamiana homologs of peanut water deficit-induced genes by virus-induced gene silencing [J]. Planta,2007,225(3):523-539.
[248]Hands P., Vosnakis N., Betts D., et al. Alternate transcripts of a floral developmental regulator have both distinct and redundant functions in Opium poppy [J]. Annals of Botany,2011,107(9):1557-1566.
[249]Kang Y. W., Kim R. N., Cho H. S., et al. Silencing of a BYPASS1 homolog results in root-independent pleiotrophic developmental defects in Nicotiana benthamiana [J]. Plant Molecular Biology,2008,68(4):423-437.
[250]Jin H., Li S., Villegas Jr A. Down-regulation of the 26S proteasome subunit RPN9 inhibits viral systemic transport and alters plant vascular development [J]. Plant Physiology,2006,142(2):651-661.
[251]Valentine T., Shaw J., Blok V. C., et al. Efficient virus-induced gene silencing in roots using a modified Tobacco rattle virus vector [J]. Plant Physiology,2004, 136(4):3999-4009.
[252]Jones L., Keining T., Eamens A., et al. Virus-induced gene silencing of Argonaute genes in Nicotiana benthamiana demonstrates that extensive systemic silencing requires Argonautel-like and Argonaute4-like genes [J]. Plant Physiology,2006,141(2):598-606.
[253]LIN Z., YIN K., WANG X., et al. Virus induced gene silencing of ATCDC5 results in accelerated cell death in Arabidopsis leaves [J]. Plant Physiology and Biochemistry,2007,45(1):87-94.
[254]Xu Qiang, Chen Ling-Ling, Ruan Xiaoan, et al. The draft genome of sweet orange (Citrus sinensis) [J]. Nature Genetics,2012,45(1):59-66.
[255]Dinesh-Kumar S. P., Burch-Smith T., Liu Y., et al. Virus-induced gene silencing (VIGS) for gene function studies in plants. [J]. Phytopathology,2007,97(7): S145-S145.
[256]Orbovic V., Soria P., Moore G. A., et al. The use of Citrus tristeza virus (CTV) containing a green fluorescent protein gene as a tool to evaluate resistance/tolerance of transgenic citrus plants [J]. Crop Protection,2011,30(5): 572-576.
[257]Ambros S., El-Mohtar C., Ruiz-Ruiz S., et al. Agroinoculation of Citrus tristeza virus causes systemic infection and symptoms in the presumed nonhost Nicotiana benthamiana [J]. Molecular Plant-Microbe Interactions,2011,24(10): 1119-1131.
[258]Lu R., Folimonov A., Shintaku M., et al. Three distinct suppressors of RNA silencing encoded by a 20-kb viral RNA genome [J]. Proceedings of the National Academy of Sciences of the United States of America,2004,101(44): 15742-15747.
[259]Vives M. C., Martin S., Ambros S., et al. Development of a full-genome cDNA clone of Citrus leaf blotch virus and infection of citrus plants [J]. Molecular Plant Pathology,2008,9(6):787-797.
[260]Renovell A., Vives M. C., Ruiz-Ruiz S., et al. The Citrus leaf blotch virus movement protein acts as silencing suppressor [J]. Virus Genes,2012,44(1): 131-140.
[261]de Assis F. F., Paguio O. R., Sherwood J. L., et al. Symptom induction by Cowpea chlorotic mottle virus on Vigna unguiculata is determined by amino acid residue 151 in the coat protein [J]. Journal of General Virology,2002,83(4): 879-883.
[262]Neeleman L., Van Der K. A. C., Bol J. F.. Role of Alfalfa mosaic virus coat protein gene in symptom formation [J]. Virology,1991,181(2):687-693.
[263]Hagiwara K., Ichiki T. U., Ogawa Y., et al. A single amino acid substitution in 126-kDa protein of Pepper mild mottle virus associates with symptom attenuation in pepper; the complete nucleotide sequence of an attenuated strain, c-1421 [J]. Archives of Virology,2002,147(4):833-840.
[264]Kay R., Chan A., Daly M., et al. Duplication of CaMV 35S promoter sequences creates a strong enhancer for plant genes [J]. Science (New York, NY),1987, 236(4806):1299-1302.
[265]Forment J., Gadea J., Huerta L., et al. Development of a citrus genome-wide EST collection and cDNA microarray as resources for genomic studies [J]. Plant Molecular Biology,2005,57(3):375-391.
[2]Wallace J. M., Drake R. J. Tatter leaf, a previously undesoribed virus effect on citrus [J]. Plant Disease Reporter,1962,46(4):211-212.
[3]Yarwood C. E. Mechanical transmission of a latent virus [J]. Phytopathology, 1963,53:1145.
[4]Fulton R. W. Mechanical transmission of tatter leaf virus from cowpea to citrus [J]. Phytopathology,1966,56(5):575.
[5]Miyakawa T., Matsui C. A bud-union abnormality of satsuma mandarin on Poncirus trifoliata rootstock in Japan [C]. Proceedings of the 7th Conference of the International Organization of Citrus Virologists. Riverside:IOCV, 1976:125-131.
[6]Miyakawa T., Tsuji M. The association of tatter leaf virus with bud-union crease of trees on trifoliate orange rootstock [C]. Proceedings of the 10th Conference of the International Organization of Citrus Virologists. Riverside:IOCV, 1988:360-364.
[7]Broadbent P., Dephoff C. M., Gilkeson C. Detection of Citrus tatter leaf virus in Australia [J]. Australasian Plant Pathology,1994,23(1):20-24.
[8]Da Graca J. V. Citrus tatter leaf virus in South African Meyer lemon [J]. Citrus Subtrop. Fruit J.,1977,529:18.
[9]Zhang T. M., Liang X. Y., Roistacher C. N. Occurrence and detection of Citrus tatter leaf virus (CTLV) in Huangyan, Zhejiang province, China [J]. Plant Disease,1988,72:543-545.
[10]Fajinmi A. A, Fajinmi O. B, Amusa N. A. An overview of citrus virus disease and its control in Nigeria [J]. Journal of Advances in Developmental Research,2011, 2(2):151-157.
[11]刘干生,刘震,刘尚泉等.柑橘碎叶病危害的调查[J].中国南方果树,2003,32(2)：22.
[12]赵小龙,张社南,刘升球等.柑橘碎叶病及其防治[J].广西园艺,2005,16(2)：32-33.
[13]Chung K., Jian W. R. Occurrence and distribution of citrus tatter leaf in Fujian, China [J].1990.
[14]Schwarz R. E. Mechanical transmission of a virus from Meyer lemon to herbaceous hosts [J]. South African Journal of Agriculture Science,1966,9: 263-264.
[15]Wallace J. M., Drake R. J. New information on symptom effects and host range of the Citrus tatter-leaf virus [J]. Plant Disease Reporter,1963,47:352-353.
[16]Roistacher C. N. A historical review of the major graft-transmissible diseases of citrus [M]. Roma (Italia):Food and Agriculture Organization of the United Nations, Regional Office for the Near East,1995.
[17]Roistacher C. N. Citrus tatter leaf virus:Further evidence for single virus complex [C]. Proceedings of the 10th Conference of the International Organization of Citrus Virologists. Riverside:IOCV,1988:353-359.
[18]Iwanami T., Kano T., Koizumi M. Pathogenic diversity of Citrus tatter leaf virus isolates [J]. Annals of the Plant Pathological Society of Japan,1991,57:74.
[19]Nishio T., Kawai A., Kato M., et al. A sap-transmissible Closterovirus in citrus imported from China and Formosa [J]. Research Bulletin of the Plant Protection Service,1982,18:11-18.
[20]陈集双,张天淼.柑橘碎叶病毒研究[J].微生物学报,1995,35(3)：173-180.
[21]Roistacher C. N., Nauer E. M., Wagner R. L. Transmissibility of Cachexia, Sweet mottle, Psorosis, Tatterleaf and Infectious variegation viruses on knife blades and its prevention [C]. Proceedings of the 8th Conference of the International Organization of Citrus Virologists. Riverside:IOCV,1980:225-229.
[22]Tanner J. D., Kunta M., Da Graca J. V., et al. Evidence of a low rate of seed transmission of Citrus tatter leaf virus in citrus [J]. Phytopathology,2011,101(6): S175-S175.
[23]Martelli G. P., Adams M. J., Kreuze J. F., et al. Family flexiviridae:A case study in virion and genome plasticity [J]. Annual Review of Phytopathology,2007,45: 73-100.
[24]King A. M. Q., Lefkowitz E., Adams M. J., et al. Virus taxonomy:Ninth Report of the International Committee on Taxonomy of Viruses [M]. London:Elsevier, 2011.
[25]Jelkmann W. Cherry virus-A-cDNA cloning of dsRNA, nucleotide-sequence analysis and serology reveal a new plant Capillovirus in sweet cherry [J]. Journal of General Virology,1995,76:2015-2024.
[26]Yoshikawa N., Sasaki E., Kato M., et al. The nucleotide sequence of Apple stem grooving Capillovirus genome [J]. Virology,1992,191(1):98-105.
[27]Komatsu K., Hirata H., Fukagawa T., et al. Infection of Capilloviruses requires subgenomic RNAs whose transcription is controlled by promoter-like sequences conserved among flexiviruses [J]. Virus Research,2012,167(1):8.
[28]Nishio T., Kawai A., Takahashi T., et al. Purification and properties of Citrus tatter leaf virus [J]. Annals of the Phytopathological Society of Japan,1989,55: 254-258.
[29]Zhao L., Hao X., Liu P., et al. Complete sequence of an Apple stem grooving virus (ASGV) isolate from China [J]. Virus Genes,2012,45(3):1-4.
[30]Magome H., Yoshikawa N., Takahashi T. Nucleotide sequence of Citrus tatter leaf virus (isolate li-23) genome [J]. Ann. Phytopathol. Soc. Jpn,1994,60:762-763.
[31]Ohira K., Namba S., Rozanov M., et al. Complete sequence of an infectious full-length cDNA clone of Citrus tatter leaf Capillovirus:Comparative sequence analysis of Capillovirus genomes [J]. Journal of General Virology,1995,76(9): 2305-2309.
[32]Magome H., Yoshikawa N., Takahashi T., et al. Molecular variability of the genomes of Capilloviruses from apple, Japanese pear, European pear, and citrus trees [J]. Phytopathology,1997,87(4):389-396.
[33]Tatineni S., Afunian M. R., Hilf M. E., et al. Molecular characterization of Citrus tatter leaf virus historically associated with Meyer lemon trees:Complete genome sequence and development of biologically active in vitro transcripts [J]. Phytopathology,2009,99(4):423-431.
[34]Bar-Joseph M., Loebenstein G., Cohen J. Partial purification of virus-like particles associated with the Citrus tristeza disease [J]. Phytopathology,1970, 60(1):75-78.
[35]Ohira K., Ito T., Kawai A., et al. Nucleotide sequence of the 3'-terminal region of Citrus tatter leaf virus RNA [J]. Virus Genes,1994,8(2):169-172.
[36]Rochon D. M., Johnston J. C. Infectious transcripts from cloned Cucumber necrosis virus cDNA:Evidence for a bifunctional subgenomic mRNA [J]. Virology,1991,181(2):656-665.
[37]Zhou H., Jackson A. O. Expression of the Barley stripe mosaic virus RNAβ "Triple Gene Block" [J]. Virology,1996,216(2):367-379.
[38]Hirata H., Yamaji Y., Komatsu K., et al. Pseudo-polyprotein translated from the full-length ORF1 of Capillovirus is important for pathogenicity, but a truncated ORF1 protein without variable and CP regions is sufficient for replication [J]. Virus Research,2010,152(1):1-9.
[39]Tatineni S., Afunian M. R., Gowda S., et al. Characterization of the 5'- and 3'-terminal subgenomic RNAs produced by a Capillovirus:Evidence for a CP subgenomic RNA [J]. Virology,2009,385(2):521-528.
[40]Calavan E. C, Christiansen D. W, Roistacher C. N. Symptoms associated with tatter leaf virus infection of Troyer citrange rootstock [J]. Plant Disease,1963, 47:971-975.
[41]Garnsey S. M, Jones J. W. Relationship of symptoms to the presence of tatter-leaf virus in several citrus hosts [C]. Proceedings of the 4th Conference of the International Organization of Citrus Virologists. Riverside:IOCV, 1968:206-212.
[42]何新华,蒋元晖,赵学源等.部分引进柑橘品种的裂皮病和碎叶病鉴定[J].四川果树,1993,21(2)：4-5.
[43]Kawai A., Nishio T. Detection of Citrus tatter leaf virus by enzyme-linked immunosorbent assay (ELISA) [J]. Annals of the Phytopathological Society of Japan,1990,56(3):342-345.
[44]Kawai A., Tsukamoto T., Namba S., et al. Citrus tatter leaf virus:A review of its properties and the development of a serological detection system [J]. Proceedings of the 13th Conference of the International Organization of Citrus Virologists. River-side:IOCV,1996:339-342.
[45]王小凤,黄谊,王文慧等.苹果茎沟病毒的提纯和检测[J].微生物学报,1997,37(4)：258-264.
[46]洪霓,王国平,于济民等.苹果茎沟病毒的分离纯化及血清学检测[J].中国农业科学,1997,30(5)：6-12.
[47]韦晓霞,吴如健.柑橘碎叶病毒的提纯[J].福建果树,1999,108(2)：17-19.
[48]Barba M., Clark M. F. Detection of strains of Apple chlorotic leafspot virus by F(ab)2-based indirect ELISA [C]. XIII International Symposium on Fruit Tree Virus Diseases 193.1985:297-304.
[49]Lovisolo O., Accotto G. P., Masenga V., et al. An isolate of Apple stem grooving virus associated with Cleopatra mandarin fruit intumescence [J]. Fitopatologia Brasileira,2003,28(1):54-58.
[50]黄训才,邓子牛,胡春华等.柑橘主要病毒病的RT-PCR法鉴定[J].湖南农业大学学报,2006,32(3)：273-276.
[51]Hailstones D. L., Bryant K. L., Broadbent P., et al. Detection of Citrus tatter leaf virus with reverse transcription-polymerase chain reaction (RT-PCR) [J]. Australasian Plant Pathology,2000,29(4):240-248.
[52]周常勇,Deborah H., Rachael C等.一种微量、快速抽提柑橘衰退病毒(ctv)核酸应用于RT-PCR扩增的方法[J].福建农业学报,2001,30(增刊)：200.
[53]唐科志,周常勇,王雪峰等.利用RT-PCR检测柑橘碎叶病毒的研究[J].植物病理学报,2005,35(6)：118-119.
[54]Ito T., Ieki H., Ozaki K. Simultaneous detection of six citrus viroids and Apple stem grooving virus from citrus plants by multiplex reverse transcription polymerase chain reaction [J]. Journal of Virological Methods,2002,106(2): 235-239.
[55]Roy A., Fayad A., Barthe G., et al. A multiplex polymerase chain reaction method for reliable, sensitive and simultaneous detection of multiple viruses in citrus trees [J]. Journal of Virological Methods,2005,129(1):47-55.
[56]肖远辉,曾继吾,张秋明等.柑橘衰退病、裂皮病和碎叶病的多重RT-PCR检测方法研究[J].植物病理学报,2007,37(1)：31-35.
[57]Heid C. A., Stevens J., Livak K. J., et al. Real time quantitative PCR [J]. Genome Research,1996,6(10):986-994.
[58]Garrido A., Chapela M. J., Roman B., et al. A new multiplex real-time PCR developed method for Salmonella spp. And listeria monocytogenes detection in food and environmental samples [J]. Food Control,2013,30(1):76-85.
[59]Xiong A. S., Jiang H. H., Zhuang J., et al. Expression and function of a modified AP2/ERF transcription factor from Brassica napus enhances cold tolerance in transgenic Arabidopsis [J]. Molecular Biotechnology,2013,53(2):198-206.
[60]郭立新,向本春,陈红运等.实时荧光RT-PCR一步法检测苹果茎沟病毒[J].植物病理学报,2006,36(1)：57-61.
[61]殷幼平,黄冠军,赵云等.柑橘溃疡病菌实时荧光定量PCR检测与应用[J].植物保护学报,2008,34(6)：607-613.
[62]刘科宏,周常勇,宋震等.运用实时荧光RT-PCR技术检测柑橘碎叶病毒[J].果树学报,2009,26(5)：748-751.
[63]何新华,将元晖.11个柑橘碎叶病毒分离株致病力强弱的比较(初报)[J].云南农业大学学报,1993,8(3)：240-241.
[64]Terauchi H, Magome H., Yoshikawa N., et al. Construction of an infectious cDNA clone of the Apple stem grooving Capillovirus (isolate li-23) genome containing a Cauliflower mosaic virus 35S RNA promoter [J]. Annals of the Phytopathological Society of Japan,1997,63.
[65]孟祥春,向旭,钟云等.砂糖橘上碎叶病毒外壳蛋白基因的克隆及序列分析 [J].果树学报,2007,24(6)：796-798.
[66]Magome H., Yoshikawa N., Takahashi T. Single-strand conformation polymorphism analysis of Apple stem grooving Capillovirus sequence variants [J]. Phytopathology,1999,89(2):136-140.
[67]Gillings M., Broadbent P., Indsto J., et al. Characterisation of isolates and strains of Citrus tristeza closterovirus using restriction analysis of the coat protein gene amplified by the polymerase chain reaction [J]. Journal of Virological Methods, 1993,44(2):305-317.
[68]魏太云,林含新,吴祖建等.应用PCR-RFLP及PCR-sscp技术研究我国水稻条纹病毒RNA4基因间隔区的变异[J].农业生物技术学报,2000,8(1)：41-44.
[69]Navarro L., Civerolo E. L, Juarez J., et al. Improvement therapy methods for citrus germplasm exchange [C]. Proceedings of the 10th Conference of the International Organization of Citrus Virologists. River-side:IOCV,1989:43
[70]何新华,蒋元晖,赵学源等.应用热处理—茎尖嫁接方法培育无碎叶病毒的柑橘[J].果树学报,1993,2：007.
[71]赵小龙,张社南,刘升球.脱除茂谷柑碎叶病毒,天草衰退病毒的试验报告[J].浙江柑橘,2006,23(3)：19-20.
[72]何新华,赵学源.柑橘碎叶病毒对八种不同来源枳砧锦橙的反应及脱毒研究[J].广西植保,1996,9(2)：4-6.
[73]Boyer J. C., Haenni A. L. Infectious transcripts and cDNA clones of RNA viruses [J]. Virology,1994,198(2):415-426.
[74]Taniguchi T., Palmieri M., Weissman C. Qβ DNA-containing hybrid plasmids giving rise to Qβ phage formation in the bacterial host [J]. Nature,1978,274 223-228.
[75]Ahlquist P., French R., Janda M., et al. Multicomponent RNA plant virus infection derived from cloned viral cDNA [J]. Proceedings of the National Academy of Sciences,1984,81(22):7066-7070.
[76]Petty I. T, Hunter B. G, Wei N., et al. Infectious Barley stripe mosaic virus RNA transcribed in vitro from full-length genomic cDNA clones [J]. Virology,1989, 171(2):342-349.
[77]Hayes R. J, Buck K. W. Infectious Cucumber mosaic virus RNA transcribed in vitro from clones obtained from cDNA amplified using the polymerase chain reaction [J]. Journal of General Virology,1990,71(11):2503-2508.
[78]Timpe U., Kuhne T. In vitro transcripts of a full-length cDNA of a naturally deleted RNA2 of Barley mild mosaic virus (BAMMV) replicate in BAMMV-infected plants [J]. Journal of General Virology,1995,76(10): 2619-2623.
[79]Meyer M., Dessens J. T.35 S promoter-driven cDNAs of Barley mild mosaic virus RNA1 and RNA2 are infectious on barley plants [J]. Journal of General Virology, 1997,78(12):3147-3151.
[80]Riechmann J. L, Lain S., Garcia JA. Infectious in vitro transcripts from a Plum poxpotyvirus cDNA clone [J]. Virology,1990,177(2):710.
[81]Sanchez F., Martnez-Herrera D., Aguilar I., et al. Infectivity of Turnip mosaic potyvirus cDNA clones and transcripts on the systemic host Arabidopsis thaliana and local lesion hosts [J]. Virus Research,1998,55(2):207-219.
[82]Kim K. S., Oh H. Y., Suranto S., et al. Infectivity of in vitro transcripts of Johnsongrass mosaic potyvirus full-length cDNA clones in maize and sorghum [J]. Archives of Virology,2003,148(3):563-574.
[83]薛朝阳,周雪平,陈青等.一种病毒侵染性全长cDNA克隆的快速构建方法[J].生物化学与生物物理学报,2000,32(3)：270-274.
[84]Dagless E. M, Shintaku M. H, Nelson R. S, et al. A CaMV 35S promoter driven cDNA clone of Tobacco mosaic virus can infect host plant tissue despite being uninfectious when manually inoculated onto leaves [J]. Archives of Virology, 1997,142(1):183-191.
[85]Yang S. J, Revers F., Souche S., et al. Construction of full-length cDNA clones of Lettuce mosaic virus (LMV) and the effects of intron-insertion on their viability in Escherichia coli and on their infectivity to plants [J]. Archives of Virology, 1998,143(12):2443-2451.
[86]Saldarelli P., Dell'Orco M., Minafra A. Infectious cDNA clones of two grapevine viruses [J]. Archives of Virology,2000,145(2):397-405.
[87]Rice C. M, Grakoui A., Galler R., et al. Transcription of infectious yellow fever RNA from full-length cDNA templates produced by in vitro ligation [J]. The New Biologist,1989,1(3):285.
[88]Forns X., Bukh J., Purcell R. H., et al. How Escherichia coli can bias the results of molecular cloning:Preferential selection of defective genomes of Hepatitis C virus during the cloning procedure [J]. Proceedings of the National Academy of Sciences,1997,94(25):13909-13914.
[89]Fakhfakh H., Vilaine F., Makni M., et al. Cell-free cloning and biolistic inoculation of an infectious cDNA of Potato virus Y [J]. Journal of General Virology,1996,77(3):519-523.
[90]Jakab G., Droz E., Brigneti G., et al. Infectious in vivo and in vitro transcripts from a full-length cDNA clone of PVY-n605, a Swiss necrotic isolate of potato virus y [J]. Journal of General Virology,1997,78(12):3141-3145.
[91]Sriburi R., Keelapang P., Duangchinda T., et al. Construction of infectious Dengue 2 virus cDNA clones using high copy number plasmid [J]. Journal of Virological Methods,2001,92(1):71-82.
[92]Skotnicki M. L, Ding S.W., Mackenzie A. M, et al. Infectious Eggplant mosaic tymovirus and Ononis yellow mosaic tymovirus from cloned cDNA [J]. Archives of Virology,1993,131(1):47-60.
[93]Sumiyoshi H., Hoke C. H., Trent D. W. Infectious Japanese encephalitis virus RNA can be synthesized from in vztro-ligated cDNA templates [J]. Journal of Virology,1992,66(9):5425-5431.
[94]Johansen I. E. Intron insertion facilitates amplification of cloned virus cDNA in Escherichia coli while biological activity is reestablished after transcription in vivo [J]. Proceedings of the National Academy of Sciences,1996,93(22): 12400-12405.
[95]Olsen B. S., Johansen I. E. Nucleotide sequence and infectious cDNA clone of the 11 isolates of Pea seed-borne mosaic potyvirus [J]. Archives of Virology, 2001,146(1):15-25.
[96]Maiss E., Timpe U., Brisske-Rode A., et al. Infectious in vivo transcripts of a Plum pox potyvirus full-length cDNA clone containing the Cauliflower mosaic virus 35S RNA promoter [J]. Journal of General Virology,1992,73(3):709-713.
[97]Vancanneyt G., Schmidt R., O'Connor-Sanchez A., et al. Construction of an intron-containing marker gene:Splicing of the intron in transgenic plants and its use in monitoring early events in Agrobacterium-mediated plant transformation [J]. Molecular and General Genetics MGG,1990,220(2):245-250.
[98]Gonzalez J. M., Penzes Z., Almazan F., et al. Stabilization of a full-length infectious cDNA clone of transmissible Gastroenteritis coronavirus by insertion of an intron [J]. Journal of Virology,2002,76(9):4655-4661.
[99]Yamshchikov V., Mishin V., Cominelli F. A new strategy in design of (+) RNA virus infectious clones enabling their stable propagation in E. Coli [J]. Virology, 2001,281(2):272-280.
[100]Polo S., Ketner G., Levis R., et al. Infectious RNA transcripts from full-length Dengue virus type 2 cDNA clones made in yeast [J]. Journal of Virology,1997, 71(7):5366-5374.
[101]MacFarlane S. A., Wallis C. V., Taylor S. C., et al. Construction and analysis of infectious transcripts synthesized from full-length cDNA clones of both genomic RNAs of Pea early browning virus [J]. Virology,1991,182(1):124-129.
[102]Flasinski S., Cassidy B. G. Potyvirus aphid transmission requires helper component and homologous coat protein for maximal efficiency [J]. Archives of Virology,1998,143(11):2159-2172.
[103]Zhang F., Huang Q., Ma W., et al. Amplification and cloning of the full-length genome of Japanese encephalitis virus by a novel long RT-PCR protocol in a cosmid vector [J]. Journal of Virological Methods,2001,96(2):171-182.
[104]Joshi A., Jeang KT. Reduction in growth temperature minimizes instability of large plasmids containing HIV-1 proviral genomes [J]. Biotechniques,1993, 14(6):880-884.
[105]Domingo E. Mutation rates and rapid evolution of RNA viruses [J]. The Evolutionary Biology of Viruses.,1994:161-184.
[106]Shimotohno K., Kodama Y., Hashimoto J., et al. Importance of 5'-terminal blocking structure to stabilize mRNA in eukaryotic protein synthesis [J]. Proceedings of the National Academy of Sciences,1977,74(7):2734-2738.
[107]Dolja V. V, McBride H. J., Carrington J.C. Tagging of plant Potyvirus replication and movement by insertion of beta-glucuronidase into the viral polyprotein [J]. Proceedings of the National Academy of Sciences,1992,89(21):10208-10212.
[108]Gal-On A., Antignus Y, Rosner A., et al. Infectious in vitro RNA transcripts derived from cloned cDNA of the Cucurbit potyvirus, Zucchini yellow mosaic virus [J]. Journal of General Virology,1991,72(11):2639-2643.
[109]Sivakumaran K., Fowler B. C., Hacker D.L. Identification of viral genes required for cell-to-cell movement of Southern bean mosaic virus [J]. Virology,1998, 252(2):376-386.
[110]Angenent G. C, Posthumus E., Bol J. F. Biological activity of transcripts synthesized in vitro from full-length and mutated DNA copies of Tobacco rattle virus RNA2 [J]. Virology,1989,173(1):68-76.
[111]Hamilton W. D. O., Baulcombe D. C. Infectious RNA produced by in vitro transcription of a full-length Tobacco rattle virus RNA-1 cDNA [J]. Journal of General Virology,1989,70(4):963-968.
[112]Huang M., Koh D. C. Y., Weng L. J., et al. Complete nucleotide sequence and genome organization of Hibiscus chlorotic ringspot virus, a new member of the genus Carmovirus:Evidence for the presence and expression of two novel open reading frames [J]. Journal of Virology,2000,74(7):3149-3155.
[113]Mayo M. A., Barker H., Robinson D. J., et al. Evidence that Potato leafroll virus RNA is positive-stranded, is linked to a small protein and does not contain polyadenylate [J]. Journal of General Virology,1982,59(1):163-167.
[114]Tobin G. J., Young D. C., Flanegan J. B. Self-catalyzed linkage of Poliovirus terminal protein VPG to Poliovirus RNA [J]. Cell,1989,59(3):511-519.
[115]Sit T. L., AbouHaidar M. G. Infectious RNA transcripts derived from cloned cDNA of Papaya mosaic virus:Effect of mutations to the capsid and polymerase proteins [J]. Journal of General Virology,1993,74(6):1133-1140.
[116]Jackson R. J., Standart N. Do the poly (A) tail and 3'untranslated region control mRNA translation? [J]. Cell,1990,62(1):15-24.
[117]Tsai C. H., Cheng C. P., Peng C. W., et al. Sufficient length of a poly (A) tail for the formation of a potential pseudoknot is required for efficient replication of Bamboo mosaic potexvirus RNA [J]. Journal of Virology,1999,73(4): 2703-2709.
[118]Chiang C. H., Yeh S. D. Infectivity assays of in vitro and in vivo transcripts of Papaya ringspot potyvirus [J]. Botanical Bulletin of Academia Sinica,1997,38: 153-163.
[119]Barnes W. M. PCR amplification of up to 35-kb DNA with high fidelity and high yield from Lambda bacteriophage templates [J]. Proceedings of the National Academy of Sciences,1994,91(6):2216-2220.
[120]Heaton L. A., Carrington J. C., Morris T. J. Turnip crinkle virus infection from RNA synthesized in vitro [J]. Virology,1989,170(1):214-218.
[121]Bujarski Jozef J., Miller W. A. Use of in vitro transcription to study gene expression and replication of spherical, positive sense RNA plant viruses [J]. Genetic Engineering with Plant Viruses,1992:115.
[122]Commandeur U., Jarausch W., Li Y., et al. cDNAs of Beet necrotic yellow vein virus RNAs 3 and 4 are rendered biologically active in a plasmid containing the Cauliflower mosaic virus 35S promoter [J]. Virology,1991,185(1):493-495.
[123]Mori M., Mise K., Kobayashi K., et al. Infectivity of plasmids containing Brome mosaic virus cDNA linked to the Cauliflower mosaic virus 35S RNA promoter [J]. Journal of General Virology,1991,72(2):243-246.
[124]Neeleman L., Van Der Vossen E. A. G., Bol J. F. Infection of tobacco with Alfalfa mosaic virus cDNAs sheds light on the early function of the coat protein [J]. Virology,1993,196(2):883-887.
[125]Zaitlin M., Palukaitis P. Advances in understanding plant viruses and virus diseases [J]. Annual Review of Phytopathology,2000,38(1):117-143.
[126]Lopez-Moya J. J., Garcia J. A. Construction of a stable and highly infectious intron-containing cDNA clone of Plum pox potyvirus and its use to infect plants by particle bombardment [J]. Virus Research,2000,68(2):99-107.
[127]Ding S. W., Rathjen J. P, Li W. X., et al. Efficient infection from cDNA clones of Cucumber mosaic cucumovirus RNAs in a new plasmid vector [J]. Journal of General Virology,1995,76(2):459-464.
[128]Weber H., Haeckel P., Pfitzner A. J. A cDNA clone of Tomato mosaic virus is infectious in plants [J]. Journal of Virology,1992,66(6):3909-3912.
[129]Gal-On A., Meiri E., Huet H., et al. Particle bombardment drastically increases the infectivity of cloned DNA of Zucchini yellow mosaic potyvirus [J]. Journal of General Virology,1995,76(12):3223-3227.
[130]Yamashita T., Iida A., Morikawa H. Evidence that more than 90% of β-glucuronidase-expressing cells after particle bombardment directly receive the foreign gene in their nucleus [J]. Plant Physiology,1991,97(2):829-831.
[131]Grimsley N., Hohn B., Hohn T., et al. "Agroinfection," an alternative route for viral infection of plants by using the Ti plasmid [J]. Proceedings of the National Academy of Sciences,1986,83(10):3282-3286.
[132]Lindbo J. A. High-efficiency protein expression in plants from agroinfection-compatible Tobacco mosaic virus expression vectors [J]. BMC Biotechnology, 2007,7(1):1-11.
[133]Ratcliff F., Martin-Hernandez A. M., Baulcombe D. C. Tobacco rattle virus as a vector for analysis of gene function by silencing [J]. Plant Journal,2001,25(2): 237-245.
[134]Shi B. J., Ding S. W., Symons R. H. Plasmid vector for cloning infectious cDNAs from plant RNA viruses:High infectivity of cDNA clones of Tomato aspermy cucumovirus [J]. Journal of General Virology,1997,78(5):1181-1185.
[135]Leiser R. M., Ziegler-Graff V., Reutenauer A., et al. Agroinfection as an alternative to insects for infecting plants with Beet western yellows luteovirus [J]. Proceedings of the National Academy of Sciences,1992,89(19):9136-9140.
[136]Grimsley N. H., Ramos C., Hein T.; et al. Meristematic tissues of maize plants are most susceptible to agroinfection with Maize streak virus [J]. Nature Biotechnology,1988,6(2):185-189.
[137]Matthew L. RNA i for plant functional genomics [J]. Comparative and Functional Genomics,2004,5(3):240-244.
[138]Van Kammen A. Virus-induced gene silencing in infected and transgenic plants [J]. Trends in Plant Science,1997,2(11):409-411.
[139]Kumagai M. H., Donson J., Della-Cioppa G., et al. Cytoplasmic inhibition of carotenoid biosynthesis with virus-derived RNA [J]. Proceedings of the National Academy of Sciences,1995,92(5):1679-1683.
[140]Godge M. R., Purkayastha A., Dasgupta I., et al. Virus-induced gene silencing for functional analysis of selected genes [J]. Plant Cell Reports,2008,27(2): 209-219.
[141]Burch-Smith T. M., Anderson J. C., Martin G. B., et al. Applications and advantages of virus-induced gene silencing for gene function studies in plants [J]. Plant Journal,2004,39(5):734-746.
[142]Baulcombe D. C. Fast forward genetics based on virus-induced gene silencing [J]. Current Opinion in Plant Biology,1999,2(2):109-113.
[143]Becker A., Lange M. Vigs-genomics goes functional [J]. Trends in Plant Science, 2010,15(1):1-4.
[144]Purkayastha A., Dasgupta I. Virus-induced gene silencing:A versatile tool for discovery of gene functions in plants [J]. Plant Physiology and Biochemistry, 2009,47(11):967-976.
[145]Senthil-Kumar M., Mysore K. S. New dimensions for VIGS in plant functional genomics [J]. Trends in Plant Science,2011,16(12):656-665.
[146]Baulcombe D. RNA silencing in plants [J]. Nature,2004,431(7006):356-363.
[147]Meyer P., Heidmann I., Niedenhof I. Differences in DNA-methylation are associated with a paramutation phenomenon in transgenic petunia [J]. The Plant Journal,2002,4(1):89-100.
[148]Llave C. Virus-derived small interfering RNAs at the core of plant-virus interactions [J]. Trends in Plant Science,2010,15(12):701-707.
[149]Donaire L., Barajas D., Martinez-Garcia B., et al. Structural and genetic requirements for the biogenesis of Tobacco rattle virus-derived small interfering RNAs [J]. Journal of Virology,2008,82(11):5167-5177.
[150]Qu F., Ye X., Morris T. J. Arabidopsis DRB4,AGO1,AGO7, and RDR6 participate in a DCL4-initiated antiviral RNA silencing pathway negatively regulated by DCL1 [J]. Proceedings of the National Academy of Sciences,2008,105(38): 14732-14737.
[151]Brodersen P., Sakvarelidze-Achard L., Bruun-Rasmussen M., et al. Widespread translational inhibition by plant miRNAs and siRNAs [J]. Science,2008, 320(5880):1185-1190.
[152]Zheng X., Zhu J., Kapoor A., et al. Role of ArabidopsisAGO6 in siRNA accumulation, DNA methylation and transcriptional gene silencing [J]. The EMBO journal,2007,26(6):1691-1701.
[153]Sasaki S., Yamagishi N., Yoshikawa N. Efficient virus-induced gene silencing in apple, pear and Japanese pear using Apple latent spherical virus vectors [J]. Plant Methods,2011,7(1):15.
[154]van der Linde K., Kastner C., Kumlehn J., et al. Systemic virus-induced gene silencing allows functional characterization of maize genes during biotrophic interaction with ustilago maydis [J]. New Phytologist,2011,189(2):471-483.
[155]Zhang C., Bradshaw J. D., Whitham S. A., et al. The development of an efficient multipurpose Bean pod mottle virus viral vector set for foreign gene expression and RNA silencing [J]. Plant Physiology,2010,153(1):52-65.
[156]Aguero J., Ruiz-Ruiz S., Vives M. D., et al. Development of viral vectors based on Citrus leaf blotch virus to express foreign proteins or analyze gene function in citrus plants [J]. Molecular Plant-Microbe Interact,2012,25(10):1326-1337.
[157]Nagamatsu A., Masuta C., Senda M., et al. Functional analysis of soybean genes involved in flavonoid biosynthesis by virus-induced gene silencing [J]. Plant Biotechnology Journal,2007,5(6):778-790.
[158]Lu H. C., Chen H. H., Tsai W. C., et al. Strategies for functional validation of genes involved in reproductive stages of orchids [J]. Plant Physiology,2007, 143(2):558-569.
[159]Kurth E. G., Peremyslov V. V., Prokhnevsky A. I., et al. Virus-derived gene expression and RNA interference vector for grapevine [J]. Journal of Virology, 2012,86(11):6002-6009.
[160]Faivre-Rampant O., Gilroy E. M., Hrubikova K., et al. Potato virus X-induced gene silencing in leaves and tubers of potato [J]. Plant Physiology,2004,134(4): 1308-1316.
[161]Gammelgard E., Mohan M., Valkonen J. P. T. Potyvirus-induced gene silencing: The dynamic process of systemic silencing and silencing suppression [J]. Journal of General Virology,2007,88(8):2337-2346.
[162]Constantin G. D., Gr(?)nlund M., Johansen I. E., et al. Virus-induced gene silencing (VIGS) as a reverse genetic tool to study development of symbiotic root nodules [J]. Molecular Plant-Microbe Interactions,2008,21(6):720-727.
[163]Naylor M., Reeves J., Cooper J. I., et al. Construction and properties of a gene-silencing vector based on Poplar mosaic virus (genus carlavirus) [J]. Journal of Virological Methods,2005,124(1-2):27-36.
[164]Vaistij F. E., Jones L. Compromised virus-induced gene silencing in RDR6-deficient plants [J]. Plant Physiology,2009,149(3):1399-1407.
[165]Varallyay E., Lichner Z., Safrany J., et al. Development of a virus induced gene silencing vector from a legumes infecting tobamovirus [J]. Acta Biologica Hungarica,2010,61(4):457-469.
[166]Hiriart J. B., Aro E. M., Lehto K. Dynamics of the VIGS-mediated chimeric silencing of the Nicotiana benthamiana chlh gene and of the Tobacco mosaic virus vector [J]. Molecular Plant-Microbe Interactions,2003,16(2):99-106.
[167]Gao Y., Zhang Y. L., Zhang X. F., et al. Development and optimization of Tobacco necrosis virus a induced gene silencing in Nicotiana benthamiana [J]. Progress in Biochemistry and Biophysics,2011,38(10):919-928.
[168]Pignatta D., Kumar P., Turina M., et al. Quantitative analysis of efficient endogenous gene silencing in Nicotiana benthamiana plants using Tomato bushy stunt virus vectors that retain the capsid protein gene [J]. Molecular Plant-Microbe Interactions,2007,20(6):609-618.
[169]Burch-Smith T. M., Schiff M., Liu Y., et al. Efficient virus-induced gene silencing in Arabidopsis [J]. Plant Physiology,2006,142(1):21-27.
[170]Fu D. Q., Zhu B. Z., Zhu H. L., et al. Enhancement of virus-induced gene silencing in tomato by low temperature and low humidity [J]. Molecules and Cells,2006,21(1):153.
[171]Ryu C. M., Anand A., Kang L., et al. Agrodrench:A novel and effective agroinoculation method for virus-induced gene silencing in roots and diverse solanaceous species [J]. The Plant Journal,2004,40(2):322-331.
[172]Fu D. Q., Zhu B. Z., Zhu H. L., et al. Virus-induced gene silencing in tomato fruit [J]. The Plant Journal,2005,43(2):299-308.
[173]Pflieger S., Blanchet S., Camborde L., et al. Efficient virus-induced gene silencing in Arabidopsis using a 'one-step'TYMV-derived vector [J]. The Plant Journal,2008,56(4):678-690.
[174]Krenz B., Wege C., Jeske H. Cell-free construction of disarmed Abutilon mosaic virus-based gene silencing vectors [J]. Journal of Virological Methods,2010, 169(1):129-137.
[175]Fofana I. B. F., Sangare A., Collier R., et al. A geminivirus-induced gene silencing system for gene function validation in cassava [J]. Plant Molecular Biology,2004,56(4):613-624.
[176]Golenberg E. M., Sather D. N., Hancock Leandria C., et al. Development of a gene silencing DNA vector derived from a broad host range Geminivirus [J]. Plant Methods,2009,5(1):1-14.
[177]Turnage M. A., Muangsan N., Peele C. G., et al. Geminivirus-based vectors for gene silencing in Arabidopsis [J]. The Plant Journal,2002,30(1):107-114.
[178]Tuttle J. R., Idris A. M, Brown J. K., et al. Geminivirus-mediated gene silencing from Cotton leaf crumple virus is enhanced by low temperature in cotton [J]. Plant Physiology,2008,148(1):41-50.
[179]Muruganantham M., Moskovitz Y., Haviv S., et al. Grapevine virusa-mediated gene silencing in Nicotiana benthamiana and Vitis vinifera [J]. Journal of Virological Methods,2009,155(2):167-74.
[180]del Rosario A. J. M., del Carmen R. G. M., Lopez M. G., et al. Virus-induced silencing of comt, pamt and kas genes results in a reduction of capsaicinoid accumulation in chili pepper fruits [J]. Planta,2008,227(3):681-695.
[181]Pandey P., Choudhury N. R., Mukherjee S. K. A geminiviral amplicon (VA) derived from Tomato leaf curl virus (ToLCV) can replicate in a wide variety of plant species and also acts as a VIGS vector [J]. Virology Journal,2009,6(1): 1-13.
[182]Gossele V., Fache I., Meulewaeter F., et al. SVISS-a novel transient gene silencing system for gene function discovery and validation in tobacco plants [J]. The Plant Journal,2002,32(5):859-866.
[183]Cai X., Wang C., Xu Y., et al. Efficient gene silencing induction in tomato by a viral satellite DNA vector [J]. Virus Research,2007,125(2):169-175.
[184]Tao X., Zhou X. A modified viral satellite DNA that suppresses gene expression in plants [J]. Plant Journal,2004,38(5):850-860.
[185]Qian Y. J., Mugiira R. B., Zhou X. P. A modified viral satellite DNA-based gene silencing vector is effective in association with Heterologous begomoviruses [J]. Virus Research,2006,118(1):136-142.
[186]Huang C. J., Xie Y., Zhou X. P. Efficient virus-induced gene silencing in plants using a modified Geminivirus DNA1 component [J]. Plant Biotechnology Journal, 2009,7(3):254-265.
[187]Ruiz M. T., Voinnet O., Baulcombe D. C. Initiation and maintenance of virus-induced gene silencing [J]. The Plant Cell Online,1998,10(6):937-946.
[188]Kjemtrup S., Sampson K. S., Peele C. G., et al. Gene silencing from plant DNA carried by a Geminivirus [J]. The Plant Journal,2002,14(1):91-100.
[189]Zhou X. P., Huang C. J. Virus-induced gene silencing using Begomovirus satellite molecules [J]. Methods Mol Biol,2012,894:57-67.
[190]Holzberg S., Brosio P., Gross C., et al. Barley stripe mosaic virus-induced gene silencing in a monocot plant [J]. Plant Journal,2002,30(3):315-327.
[191]Scofield S. R., Brandt A. S. Virus-induced gene silencing in hexaploid wheat using Barley stripe mosaic virus vectors [J]. Methods in molecular biology,2012, 894:93-112.
[192]Scofield S. R., Huang L., Brandt A. S., et al. Development of a virus-induced gene-silencing system for hexaploid wheat and its use in functional analysis of the lr21-mediated leaf rust resistance pathway [J]. Plant Physiology,2005, 138(4):2165-2173.
[193]Constantin G. D., Krath B. N., MacFarlane S. A., et al. Virus-induced gene silencing as a tool for functional genomics in a legume species [J]. Plant Journal, 2004,40(4):622-631.
[194]Zhang C., Ghabrial S. A. Development of Bean pod mottle virus-based vectors for stable protein expression and sequence-specific virus-induced gene silencing in soybean [J]. Virology,2006,344(2):401-11.
[195]Zhang C., Yang C., Whitham S. A., et al. Development and use of an efficient DNA-based viral gene silencing vector for soybean [J]. Molecular Plant-Microbe Interactions,2009,22(2):123-131.
[196]Ding X. S., Schneider W. L., Chaluvadi S. R., et al. Characterization of a brome mosaic virus strain and its use as a vector for gene silencing in monocotyledonous hosts [J]. Molecular Plant-Microbe Interact,2006,19(11): 1229-1239.
[197]Idris A. M., Ktenz B., Tuttle J. R., et al. Construction of a virus-induced gene silencing (VIGS)vector for cotton using Cotton leaf crumple virus and a fragment of the cotton phytoene desaturase [J]. Phytopathology,2008,98(6):S70-S70.
[198]Jones A. L., Thomas C. L., Maule A. J. De novo methylation and co-suppression induced by a cytoplasmically replicating plant RNA virus [J]. The EMBO Journal, 1998,17(21):6385-6393.
[199]Kanazawa A., Inaba J., Shimura H., et al. Virus-mediated efficient induction of epigenetic modifications of endogenous genes with phenotypic changes in plants [J]. The Plant Journal,2011,65(1):156-168.
[200]Sonoda S., Nishiguchi M. Delayed activation of post-transcriptional gene silencing and de novo transgene methylation in plants with the coat protein gene of Sweet potato feathery mottle potyvirus [J]. Plant Science,2000,156(2): 137-144.
[201]Vainstein A., Marton I., Zuker A., et al. Permanent genome modifications in plant cells by transient viral vectors [J]. Trends in Biotechnology,2011,29(8): 363-369.
[202]Kim S., Kim J. S. Targeted genome engineering via zinc finger nucleases [J]. Plant Biotechnology Reports,2011,5(1):9-17.
[203]Meng Y., Moscou M. J., Wise R. P. Blufensinl negatively impacts basal defense in response to Barley powdery mildew [J]. Plant Physiology,2009,149(1): 271-285.
[204]Liu Y., Schiff M., Dinesh-Kumar S. P. Virus-induced gene silencing in tomato [J]. Plant Journal,2002,31(6):777-786.
[205]Dong Y., Burch-Smith T. M., Liu Y., et al. A ligation-independent cloning Tobacco rattle virus vector for high-throughput virus-induced gene silencing identifies roles for NbMADS4-1 and-2 in floral development [J]. Plant Physiology, 2007,145(4):1161-1170.
[206]Tang Y, Wang F., Zhao J., et al. Virus-based microRNA expression for gene functional analysis in plants [J]. Plant Physiology,2010,153(2):632-641.
[207]Lacomme C. Milestones in the development and applications of plant virus vector as gene silencing platforms [J]. Current Topics in Microbiology and Immunology, 2011,10.1007/82_2011_186.
[208]Valentine T. A., Randall E., Wypijewski K., et al. Delivery of macromolecules to plant parasitic nematodes using a Tobacco rattle virus vector [J]. Plant Biotechnology Journal,2007,5(6):827-834.
[209]Dubreuil G., Magliano M., Dubrana M. P., et al. Tobacco rattle virus mediates gene silencing in a plant parasitic root-knot nematode [J]. Journal of Experimental Botany,2009,60(14):4041-4050.
[210]Hein I., Barciszewska-Pacak M., Hrubikova K., et al. Virus-induced gene silencing-based functional characterization of genes associated with powdery mildew resistance in barley [J]. Plant Physiology,2005,138(4):2155-2164.
[211]Goodin M. M., Zaitlin D., Naidu R. A., et al. Nicotiana benthamiana:Its history and future as a model for plant-pathogen interactions [J]. Molecular Plant-Microbe Interactions,2008,21(8):1015-1026.
[212]Yang S. J., Carter S. A., Cole A. B., et al. A natural variant of a host RNA-dependent RNA polymerase is associated with increased susceptibility to viruses by Nicotiana benthamiana [J]. Proceedings of the National Academy of Sciences of the United States of America,2004,101(16):6297-6302.
[213]Thomas C. L., Jones L., Baulcombe D. C., et al. Size constraints for targeting post-transcriptional gene silencing and for RNA-directed methylation in Nicotiana benthamiana using a Potato virus X vector [J]. The Plant Journal,2001, 25(4):417-425.
[214]Xu P., Zhang Y., Kang L., et al. Computational estimation and experimental verification of off-target silencing during posttranscriptional gene silencing in plants [J]. Plant Physiology,2006,142(2):429-440.
[215]Senthil-Kumar M., Mysore K. S. Caveat of RNAi in plants:The off-target effect [J]. RNAi and Plant Gene Function Analysis,2011:13-25.
[216]Senthil-Kumar M., Hema R., Anand A., et al. A systematic study to determine the extent of gene silencing in Nicotiana benthamiana and other Solanaceae Species when heterologous gene sequences are used for virus-induced gene silencing [J]. New Phytologist,2007,176(4):782-791.
[217]Jacob S. S., Vanitharani R., Karthikeyan A. S., et al. Mungbean yellow mosaic virus-Vi agroinfection by codelivery of DNA A and DNA B from one Agrobacterium strain [J]. Plant disease,2003,87(3):247-251.
[218]Liu E., Page J. E. Optimized cDNA libraries for virus-induced gene silencing (VIGS) using Tobacco rattle virus [J]. Plant Methods,2008,4(5):1-13.
[219]Rodrigo G., Carrera J., Jaramillo A., et al. Optimal viral strategies for bypassing RNA silencing [J]. Journal of The Royal Society Interface,2011,8(55):257-268.
[220]Lacomme C, Hrubikova K. Enhancement of virus-induced gene silencing through viral-based production of inverted-repeats [J]. The Plant Journal, 2003,34(4):543-553.
[221]Molnar A., Csorba T., Lakatos L., et al. Plant virus-derived small interfering RNAs originate predominantly from highly structured single-stranded viral RNAs [J]. Journal of Virology,2005,79(12):7812-7818.
[222]Szittya G., Silhavy D., Molnar A., et al. Low temperature inhibits RNA silencing-mediated defence by the control of siRNA generation [J]. The EMBO Journal,2003,22(3):633-640.
[223]Orzaez D., Mirabel S., Wieland W. H., et al. Agroinjection of tomato fruits. A tool for rapid functional analysis of transgenes directly in fruit [J]. Plant Physiology,2006,140(1):3-11.
[224]Romero I., Tikunov Y., Bovy A. Virus-induced gene silencing in detached tomatoes and biochemical effects of phytoene desaturase gene silencing [J]. Journal of Plant Physiology,2011,168(10):1129-1135.
[225]Unver T., Budak H. Virus-induced gene silencing, a post transcriptional gene silencing method [J]. International Journal of Plant Genomics,2009:1986-1980.
[226]Lucioli A., Noris E., Brunetti A., et al. Tomato yellow leaf curl Sardinia virus rep-derived resistance to homologous and heterologous Geminiviruses occurs by different mechanisms and is overcome if virus-mediated transgene silencing is activated [J]. Journal of Virology,2003,77(12):6785-6798.
[227]Lu R., Malcuit I., Moffett P., et al. High throughput virus-induced gene silencing implicates heat shock protein 90 in plant disease resistance [J]. The EMBO Journal,2003,22(21):5690-5699.
[228]Chen J. C., Jiang C. Z., Gookin T. E., et al. Chalcone synthase as a reporter in virus-induced gene silencing studies of flower senescence [J]. Plant Molecular Biology,2004,55(4):521-530.
[229]Huang C.J., Qian Y.J., Li Z.H., et al. Virus-induced gene silencing and its application in plant functional genomics [J]. Science China Life Sciences,2012, 55(2):99-108.
[230]Liu Y., Schiff M., Czymmek K., et al. Autophagy regulates programmed cell death during the plant innate immune response [J]. Cell,2005,121(4):567-577'.
[231]Caplan J.L., Mamillapalli P., Burch-Smith T.M., et al. Chloroplastic protein NRIP1 mediates innate immune receptor recognition of a viral effector [J]. Cell, 2008,132(3):449-462.
[232]Gronlund M., Olsen A., Johansen E. I., et al. Protocol:Using virus-induced gene silencing to study the arbuscular mycorrhizal symbiosis in Pisum sativum [J]. Plant Methods,2010,6(1):28.
[233]Zhu X., Dinesh-Kumar S. P. Virus-induced gene silencing as a tool to identify host genes affecting viral pathogenicity [J]. Plant Virology Protocols,2008,451: 641-648.
[234]Cloutier S., McCallum B. D., Loutre C., et al. Leaf rust resistance gene IR1, isolated from bread wheat (Triticum aestivum 1.) is a member of the large psr567 gene family [J]. Plant Molecular Biology,2007,65(1):93-106.
[235]Borras-Hidalgo O., Thomma B. P. H. J., Collazo C., et al. EIL2 transcription factor and glutathione synthetase are required for defense of tobacco against Tobacco blue mold [J]. Molecular Plant-Microbe Interactions,2006,19(4): 399-406.
[236]Ekengren S.K., Liu Y., Schiff M., et al. Two MAPK cascades, NPR1, and TGA transcription factors play a role in Pto-mediated disease resistance in tomato [J]. The Plant Journal,2003,36(6):905-917.
[237]Sharma P. C., Ito A., Shimizu T., et al. Virus-induced silencing of WIPK and SIPK genes reduces resistance to a bacterial pathogen, but has no effect on the INF1-induced hypersensitive response (HR) in Nicotiana benthamiana [J]. Molecular Genetics and Genomics,2003,269(5):583-591.
[238]Maimbo M., Ohnishi K., Hikichi Y., et al. S-glycoprotein-like protein regulates defense responses in Nicotiana plants against Ralstonia solanacearum [J]. Plant Physiology,2010,152(4):2023-2035.
[239]Lee D. H., Choi H. W., Hwang B. K. The pepper E3 ubiquitin ligase RING1 gene, CaRINGl, is required for cell death and the salicylic acid-dependent defense response [J]. Plant Physiology,2011,156(4).
[240]Melech-Bonfil S., Sessa G. The SLMKK2 and SLMPK2 genes play a role in tomato disease resistance to Xanthomonas campestris pv. Vesicatoria [J]. Plant Signaling & Behavior,2011,6(1):154-156.
[241]Mantelin S., Peng H. C., Li B., et al. The receptor-like kinase SLSERK1 is required for Mi-1-mediated resistance to potato aphids in tomato [J]. The Plant Journal,2011,67(3):459-471.
[242]Lou Y., Baldwin I. T. Silencing of a germin-like gene in Nicotiana attenuata improves performance of native herbivores [J]. Plant Physiology,2006,140(3): 1126-1136.
[243]Heinrich M., Baldwin I. T., Wu J. Two mitogen-activated protein kinase kinases, MKK1 and MEK2, are involved in wounding-and specialist Lepidopteran herbivore manduca sexta-induced responses in nicotiana attenuata [J]. Journal of Experimental Botany,2011,62(12):4355-4365.
[244]Guo Y., Huang C., Xie Y., et al. A tomato glutaredoxin gene SLGX1 regulates plant responses to oxidative, drought and salt stresses [J]. Planta,2010,232(6): 1499-1509.
[245]Re D. A., Dezar C. A., Chan R. L., et al. Nicotiana attenuata NaHD20 plays a role in leaf ABA accumulation during water stress, benzylacetone emission from flowers, and the timing of bolting and flower transitions [J]. Journal of Experimental Botany,2011,62(1):155-166.
[246]Lee S. C., Choi D. S., Hwang I. S., et al. The pepper oxidoreductase CAOXR1 interacts with the transcription factor caravl and is required for salt and osmotic stress tolerance [J]. Plant Molecular Biology,2010,73(4):409-424.
[247]Senthil-Kumar M., Govind G., Kang L., et al. Functional characterization of Nicotiana benthamiana homologs of peanut water deficit-induced genes by virus-induced gene silencing [J]. Planta,2007,225(3):523-539.
[248]Hands P., Vosnakis N., Betts D., et al. Alternate transcripts of a floral developmental regulator have both distinct and redundant functions in Opium poppy [J]. Annals of Botany,2011,107(9):1557-1566.
[249]Kang Y. W., Kim R. N., Cho H. S., et al. Silencing of a BYPASS1 homolog results in root-independent pleiotrophic developmental defects in Nicotiana benthamiana [J]. Plant Molecular Biology,2008,68(4):423-437.
[250]Jin H., Li S., Villegas Jr A. Down-regulation of the 26S proteasome subunit RPN9 inhibits viral systemic transport and alters plant vascular development [J]. Plant Physiology,2006,142(2):651-661.
[251]Valentine T., Shaw J., Blok V. C., et al. Efficient virus-induced gene silencing in roots using a modified Tobacco rattle virus vector [J]. Plant Physiology,2004, 136(4):3999-4009.
[252]Jones L., Keining T., Eamens A., et al. Virus-induced gene silencing of Argonaute genes in Nicotiana benthamiana demonstrates that extensive systemic silencing requires Argonautel-like and Argonaute4-like genes [J]. Plant Physiology,2006,141(2):598-606.
[253]LIN Z., YIN K., WANG X., et al. Virus induced gene silencing of ATCDC5 results in accelerated cell death in Arabidopsis leaves [J]. Plant Physiology and Biochemistry,2007,45(1):87-94.
[254]Xu Qiang, Chen Ling-Ling, Ruan Xiaoan, et al. The draft genome of sweet orange (Citrus sinensis) [J]. Nature Genetics,2012,45(1):59-66.
[255]Dinesh-Kumar S. P., Burch-Smith T., Liu Y., et al. Virus-induced gene silencing (VIGS) for gene function studies in plants. [J]. Phytopathology,2007,97(7): S145-S145.
[256]Orbovic V., Soria P., Moore G. A., et al. The use of Citrus tristeza virus (CTV) containing a green fluorescent protein gene as a tool to evaluate resistance/tolerance of transgenic citrus plants [J]. Crop Protection,2011,30(5): 572-576.
[257]Ambros S., El-Mohtar C., Ruiz-Ruiz S., et al. Agroinoculation of Citrus tristeza virus causes systemic infection and symptoms in the presumed nonhost Nicotiana benthamiana [J]. Molecular Plant-Microbe Interactions,2011,24(10): 1119-1131.
[258]Lu R., Folimonov A., Shintaku M., et al. Three distinct suppressors of RNA silencing encoded by a 20-kb viral RNA genome [J]. Proceedings of the National Academy of Sciences of the United States of America,2004,101(44): 15742-15747.
[259]Vives M. C., Martin S., Ambros S., et al. Development of a full-genome cDNA clone of Citrus leaf blotch virus and infection of citrus plants [J]. Molecular Plant Pathology,2008,9(6):787-797.
[260]Renovell A., Vives M. C., Ruiz-Ruiz S., et al. The Citrus leaf blotch virus movement protein acts as silencing suppressor [J]. Virus Genes,2012,44(1): 131-140.
[261]de Assis F. F., Paguio O. R., Sherwood J. L., et al. Symptom induction by Cowpea chlorotic mottle virus on Vigna unguiculata is determined by amino acid residue 151 in the coat protein [J]. Journal of General Virology,2002,83(4): 879-883.
[262]Neeleman L., Van Der K. A. C., Bol J. F.. Role of Alfalfa mosaic virus coat protein gene in symptom formation [J]. Virology,1991,181(2):687-693.
[263]Hagiwara K., Ichiki T. U., Ogawa Y., et al. A single amino acid substitution in 126-kDa protein of Pepper mild mottle virus associates with symptom attenuation in pepper; the complete nucleotide sequence of an attenuated strain, c-1421 [J]. Archives of Virology,2002,147(4):833-840.
[264]Kay R., Chan A., Daly M., et al. Duplication of CaMV 35S promoter sequences creates a strong enhancer for plant genes [J]. Science (New York, NY),1987, 236(4806):1299-1302.
[265]Forment J., Gadea J., Huerta L., et al. Development of a citrus genome-wide EST collection and cDNA microarray as resources for genomic studies [J]. Plant Molecular Biology,2005,57(3):375-391.