Expression of the CMV-CP Gene in Synechocystis
 6803 Affects Cyanobacterial Photosynthesis

ABSTRACTThe effect of large-scale synonymous substitutions in a small icosahedral, single-stranded RNA viral genome on virulence, viral titer, and protein evolution were analyzed. The coat protein (CP) gene of the Fny stain of cucumber mosaic virus (CMV) was modified. We created four CP mutants in which all the codons of nine amino acids in the 5′ or 3′ half of the CP gene were replaced by either the most frequently or the least frequently used synonymous codons in monocot plants. When the dicot host (Nicotiana benthamiana) was inoculated with these four CP mutants, viral RNA titers in uninoculated symptomatic leaves decreased, while all mutants eventually showed mosaic symptoms similar to those for the wild type. The codon adaptation index of these four CP mutants against dicot genes was similar to those of the wild-type CP gene, indicating that the reduction of viral RNA titer was due to deleterious changes of the secondary structure of RNAs 3 and 4. When two 5′ mutants were serially passaged inN. benthamiana, viral RNA titers were rapidly restored but competitive fitness remained decreased. Although no nucleic acid changes were observed in the passaged wild-type CMV, one to three amino acid changes were observed in the synonymously mutated CP of each passaged virus, which were involved in recovery of viral RNA titer of 5′ mutants. Thus, we demonstrated that deleterious effects of the large-scale synonymous substitutions in the RNA viral genome facilitated the rapid amino acid mutation(s) in the CP to restore the viral RNA titer.IMPORTANCERecently, it has been known that synonymous substitutions in RNA virus genes affect viral pathogenicity and competitive fitness by alteration of global or local RNA secondary structure of the viral genome. We confirmed that large-scale synonymous substitutions in the CP gene of CMV resulted in decreased viral RNA titer. Importantly, when viral evolution was stimulated by serial-passage inoculation, viral RNA titer was rapidly restored, concurrent with a few amino acid changes in the CP. This novel finding indicates that the deleterious effects of large-scale nucleic acid mutations on viral RNA secondary structure are readily tolerated by structural changes in the CP, demonstrating a novel part of the adaptive evolution of an RNA viral genome. In addition, our experimental system for serial inoculation of large-scale synonymous mutants could uncover a role for new amino acid residues in the viral protein that have not been observed in the wild-type virus strains.

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Identification, Characterization, and Molecular Detection of Alfalfa mosaic virus in Potato

Phytopathology ◽

10.1094/phyto-96-1237 ◽

2006 ◽

Vol 96 (11) ◽

pp. 1237-1242 ◽

Cited By ~ 34

Author(s):

H. Xu ◽

J. Nie

Keyword(s):

Mosaic Virus ◽

Gene Sequence ◽

Alfalfa Mosaic Virus ◽

Enzyme Linked Immunosorbent Assay ◽

Polymorphism Analysis ◽

Rt Pcr ◽

Potato Leaf ◽

Simple Method ◽

Cp Gene ◽

Nucleotide Sequence Alignment

Alfalfa mosaic virus (AMV) was detected in potato fields in several provinces in Canada and characterized by bioassay, enzyme-linked immunosorbent assay, and reverse-transcription polymerase chain reaction (RT-PCR). The identity of eight Canadian potato AMV isolates was confirmed by sequence analysis of their coat protein (CP) gene. Sequence and phylogenetic analysis indicated that these eight AMV potato isolates fell into one strain group, whereas a slight difference between Ca175 and the other Canadian AMV isolates was revealed. The Canadian AMV isolates, except Ca175, clustered together among other strains based on alignment of the CP gene sequence. To detect the virus, a pair of primers, AMV-F and AMV-R, specific to the AMV CP gene, was designed based on the nucleotide sequence alignment of known AMV strains. Evaluations showed that RT-PCR using this primer set was specific and sensitive for detecting AMV in potato leaf and tuber samples. AMV RNAs were easily detected in composite samples of 400 to 800 potato leaves or 200 to 400 tubers. Restriction analysis of PCR amplicons with SacI was a simple method for the confirmation of PCR tests. Thus, RT-PCR followed by restriction fragment length polymorphism analysis may be a useful approach for screening potato samples on a large scale for the presence of AMV.

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Localization of the Ceruloplasmin (CP) Gene to the q32-q33 Bands of Chromosome 13 in Pigs by in Situ Hybridization

Hereditas ◽

10.1111/j.1601-5223.1993.00007.x ◽

2004 ◽

Vol 119 (1) ◽

pp. 7-10 ◽

Cited By ~ 3

Author(s):

Renuka Chaudhary ◽

Bhanu P. Chowdhary ◽

Ingrid Harbitz ◽

Ingemar Gustavsson

Keyword(s):

In Situ Hybridization ◽

Chromosome 13 ◽

Cp Gene

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Prevalence and Molecular Characterization Coat Protein Gene of Tobacco streak virus Causing Peanut Stem Necrosis Disease in Coastal and Rayalaseema Districts of Andhra Pradesh, South-India

Legume Research - An International Journal ◽

10.18805/lr-4515 ◽

2021 ◽

Author(s):

K. Saratbabu ◽

K. Vemana ◽

A.K. Patibanda ◽

B. Sreekanth ◽

V. Srinivasa Rao

Keyword(s):

Coat Protein ◽

Molecular Characterization ◽

Andhra Pradesh ◽

Disease Incidence ◽

Tobacco Streak Virus ◽

Enzyme Linked Immunosorbent Assay ◽

Streak Virus ◽

Rt Pcr ◽

Cp Gene ◽

Stem Necrosis

Background: Peanut stem necrosis disease (PSND) caused by Tobacco streak virus (TSV) is a major constraint for groundnut production in Andhra Pradesh (A.P.). However, studies on prevalence and spread of the disease confined to only few districts of A.P. with this background current study focused on incidence and spread of the disease in entire state of A.P. Further an isolate of TSV occurring in A.P. characterized on the basis of genetic features by comparing with other TSV isolates originated from different hosts and locations from world.Methods: Roving survey was conducted during kharif 2017-18 in groundnut growing districts of Andhra Pradesh (A.P.) for peanut stem necrosis disease incidence. Groundnut plants showing PSND symptoms were collected and tested with direct antigen coating enzyme linked immunosorbent assay (DAC-ELISA). Groundnut samples found positive by ELISA once again tested by reverse transcription polymerase chain reaction (RT-PCR). The representative TSV-GN-INDVP groundnut isolate from Prakasham district was maintained on cowpea seedlings by standard sap inoculation method in glasshouse for further molecular characterization. The Phylogenetic tree for coat protein (CP) gene was constructed using aligned sequences with 1000 bootstrap replicates following neighbor-joining phylogeny.Result: Thirty-eight (52.7%) of seventy-two groundnut samples collected from different locations in A.P were given positive reaction to TSV by DAC-ELISA. For the first time, PSND incidence observed in coastal districts (Krishna, Guntur, Sri Pottisriramulu Nellore, Prakasham) of A.P. Maximum PSND incidence recorded from Bathalapalli (22.2%) and the minimum incidence in Mulakalacheruvu (4.1%). The coat protein (CP) gene of TSV-GN-INDVP groundnut isolate was amplified by RT-PCR and it shared maximum per cent nucleotide identity (97.51-98.62%) with TSV isolates from groundnut and other different crops reported in India. All Indian isolates cluster together irrespective of crop and location based on the phylogenetic analysis.

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Production of Antibodies to Citrus Tristeza Virus in Transgenic Citrus

10.32747/1995.7613018.bard ◽

1995 ◽

Author(s):

Richard Lee ◽

Moshe Bar-Joseph ◽

K.S. Derrick ◽

Aliza Vardi ◽

Roland Brlansky ◽

...

Keyword(s):

Citrus Tristeza Virus ◽

Virus Disease ◽

Biological Activities ◽

Cp Gene ◽

Single Domain Antibodies ◽

Tristeza Virus ◽

Stem Pitting ◽

Citrus Tristeza

Citrus tristeza virus (CTV) is the most important virus disease of citrus in the world. CTV causes death of trees on sour orange rootstock and/or stem pitting of scions regardless of rootstock which results in trees of low vigor, reduced yield with reduction in size and quality of fruit. The purpose of this project was to produce monoclonal antibodies (MABs) to CTV coat protein (CP), develop single domain antibodies (dAbs) or Fab fragments which neutralize the infection by binding to the virus, and to produce transformed plants which express the dAbs. The objectives of this research have been met and putative transgenic tobacco and citrus plants have been developed. These putative transgenic plants are presently undergoing evaluation to determine the level of dAbs expression and to determine their resistance to CTV. Additionally, the CTV genome has been sequenced and the CP gene of several biologically characterized CTV strains molecular characterized. This has indicated a correlation between CP sequence homology and biological activity, and the finding of DI RNAs associated with some CTV strains. Several MABs have been produced which enable broad spectrum identification of CTV strains while other MABs enable differentiation between mild and severe strains. The use of selected MAbs and determination of the CP gene sequence has enabled predictions of biological activities of unknown CTV isolates. The epitopes of two MABs, one reacting selectively with severe CTV strains and the other reacting with all strains, have been characterized at the molecular level.

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Comparative reactions of recombinant papaya ringspot viruses with chimeric coat protein (CP) genes and wild-type viruses on CP-transgenic papaya

Journal of General Virology ◽

10.1099/0022-1317-82-11-2827 ◽

2001 ◽

Vol 82 (11) ◽

pp. 2827-2836 ◽

Cited By ~ 32

Author(s):

Chu-Hui Chiang ◽

Ju-Jung Wang ◽

Fuh-Jyh Jan ◽

Shyi-Dong Yeh ◽

Dennis Gonsalves

Keyword(s):

Coat Protein ◽

Sequence Similarity ◽

Transcriptional Gene Silencing ◽

Papaya Ringspot Virus ◽

Wild Type ◽

Coding Region ◽

Transgenic Papaya ◽

Cp Gene ◽

Post Transcriptional Gene Silencing

Transgenic papaya cultivars SunUp and Rainbow express the coat protein (CP) gene of the mild mutant of papaya ringspot virus (PRSV) HA. Both cultivars are resistant to PRSV HA and other Hawaii isolates through homology-dependent resistance via post-transcriptional gene silencing. However, Rainbow, which is hemizygous for the CP gene, is susceptible to PRSV isolates from outside Hawaii, while the CP-homozygous SunUp is resistant to most isolates but susceptible to the YK isolate from Taiwan. To investigate the role of CP sequence similarity in overcoming the resistance of Rainbow, PRSV HA recombinants with various CP segments of the YK isolate were constructed and evaluated on Rainbow, SunUp and non-transgenic papaya. Non-transgenic papaya were severely infected by all recombinants, but Rainbow plants developed a variety of symptoms. On Rainbow, a recombinant with the entire CP gene of YK caused severe symptoms, while recombinants with only partial YK CP sequences produced a range of milder symptoms. Interestingly, a recombinant with a YK segment from the 5′ region of the CP gene caused very mild, transient symptoms, whereas recombinants with YK segments from the middle and 3′ parts of the CP gene caused prominent and lasting symptoms. SunUp was resistant to all but two recombinants, which contained the entire CP gene or the central and 3′-end regions of the CP gene and the 3′ non-coding region of YK, and the resulting symptoms were mild. It is concluded that the position of the heterologous sequences in the recombinants influences their pathogenicity on Rainbow.

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First Report of a Tobamovirus in Dieffenbachia and Impatiens

Plant Disease ◽

10.1094/pdis.2000.84.6.707a ◽

2000 ◽

Vol 84 (6) ◽

pp. 707-707 ◽

Cited By ~ 2

Author(s):

E. B. Rivas ◽

E. F. Pezani ◽

M. A. V. Alexandre ◽

L. M. L. Duarte

Keyword(s):

Ornamental Plants ◽

Start Codon ◽

Enzyme Linked Immunosorbent Assay ◽

Mechanical Transmission ◽

Reaction Products ◽

Cp Gene ◽

Gomphrena Globosa ◽

Leaf Deformation ◽

Antisense Primer ◽

Impatiens Hawkeri

Tobamoviruses were detected in two ornamental plants, Dieffenbachia picta (Araceae) and Impatiens hawkeri (Balsaminaceae), from different counties in São Paulo State, Brazil. Symptoms were chlorotic spots and rings in D. picta and mosaic, blistering, and leaf deformation in I. hawkeri. Mechanical transmission from both species induced different kinds and intensities of symptoms in the same experimental hosts (Balsaminaceae, Chenopodiaceae, and Solanaceae), except Gomphrena globosa, which was infected only by the isolate from D. picta. The viruses did not infect Cucurbitaceae and Fabaceae. Indirect enzyme-linked immunosorbent assay performed with extracts from infected Nicotiana tabacum ‘White Burley’ and antisera against Cucumber green mottle, mosaic, Frangipani mosaic, Odontoglossum ringspot, Ribgrass mosaic, Tobacco mosaic (TMV), Tomato mosaic, Turnip vein clearing, and Youcai mosaic viruses (genus To-bamovirus) was positive only for TMV. Furthermore, the viruses isolated from D. picta and I. hawkeri cross-reacted with their heterologous antisera. Two sense primers for regions ≍200 and 90 nt upstream of the start codon and an antisense primer ≍60 nt downstream of the terminal codon of the coat protein (CP) gene were designed for two amplification assays. Migrating fragments the same size as the reverse-transcription polymerase chain reaction products from the TMV type strain (479 and 800 bp with internal and external primers, respectively) were produced. The CP gene sequence will allow comparison and identification of the two viruses isolated from D. picta and I. hawkeri.

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Cucurbit leaf curl virus, a New Whitefly Transmitted Geminivirus in Arizona, Texas, and Mexico

Plant Disease ◽

10.1094/pdis.2000.84.7.809a ◽

2000 ◽

Vol 84 (7) ◽

pp. 809-809 ◽

Cited By ~ 19

Author(s):

J. K. Brown ◽

A. M. Idris ◽

M. W. Olsen ◽

M. E. Miller ◽

T. Isakeit ◽

...

Keyword(s):

Binding Site ◽

Field Isolate ◽

Bipartite Begomovirus ◽

Degenerate Primers ◽

Field Isolates ◽

Squash Leaf Curl Virus ◽

Cp Gene ◽

Honeydew Melon ◽

Leaf Curl Virus ◽

Leaf Curl

In 1998 to 1999, geminivirus-like symptoms were observed in whitefly-infested pumpkin, honeydew melon, and muskmelon in Arizona and Texas and in Coahuilla, Mexico (MX), respectively. Plants exhibited leaf curl and/or mottling, reminiscent of symptoms caused by Squash leaf curl virus (SLCV-WAZ) described from Arizona in 1981 (2). The isolate from Arizona pumpkin fields was experimentally transmitted to pumpkin seedlings by the “B type” of Bemisia tabaci (Genn.), and symptoms were indistinguishable from those observed in infected fields. Samples from AZ, MX, and TX were assessed for begomovirus presence by polymerase chain reaction (PCR) using degenerate primers that amplify a contiguous fragment containing the viral coat protein (Cp) gene and common region (CR) of the A component (CR-A) (~2,100 bp) and a fragment containing the CR of the B component (CR-B) (~1,100 bp). One to four isolates from each location were examined by PCR using both primer pairs, and at least three amplicons per isolate were cloned and their sequences determined. Alignment of viral Cp nucleotide (nt) sequences revealed that AZ [AF256199], MX, and TX field isolates shared 98.7 to 100% sequence identity, but were only 84.5 to 85.6% identical to the Cp gene of SLCV-extended (SLCV-E) [M38183] and SLCV-restricted (SLCV-R) (S. G. Lazarowitz, unpublished), respectively, suggesting a new, previously undescribed begomoviral species (3). Further, the Cp nt sequence of the three field isolates was 6 nt shorter than SLCV-E, SLCV-WAZ [AF256203], and SLCV-R Cp sequences. The CR-A [AF256200] and CR-B [AF256201] sequences (179 nt, each) of field isolates, including the theoretical Rep binding element, GGTGT, were 100% identical. Although the Rep binding site is identical among field isolates, SLCV-E, SLCV-R, and SLCV-WAZ, the field isolate CR sequence shared only 64.2, 67.5, and 66.9% overall identity with CR-A SLCV-E, SLCV-R [M63155], and SLCV-WAZ [AF256202], respectively. Prior to 1998 to 1999, SLCV-WAZ was the only New World begomovirus of cucurbits known to infect both melon (Cucumis) and pumpkin (Cucurbita) (1). Therefore, SLCV was initially suspected as the causal agent. However, here we provide evidence for a new, previously undescribed bipartite begomovirus of cucurbits in AZ, MX, and TX that is herein provisionally designated Cucurbit leaf curl virus (CuLCV). Prediction of its closest begomovirus relatives by Cp nt sequence and Rep binding site comparisons suggest that CuLCV is a new member of the SLCV lineage, also containing Bean calico mosaic virus, Cabbage leaf curl virus, SLCV-E, and Texas pepper virus-TAM. References: (1) J. K. Brown and M. R. Nelson. Phytopathology 74:1136, 1984. (2) J. K. Brown and M. R. Nelson. Ann. Appl. Biol. 115:243, 1986. (3) M. A. Mayo and C. R. Pringle. J. Gen. Virol. 97:649, 1998.

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