scholarly journals Robust Response to Plum pox virus Infection via Plant Biotechnology

Genes ◽  
2021 ◽  
Vol 12 (6) ◽  
pp. 816
Author(s):  
Michel Ravelonandro ◽  
Pascal Briard ◽  
Ralph Scorza ◽  
Ann Callahan ◽  
Ioan Zagrai ◽  
...  
Keyword(s):  
Genetic Resource ◽  
Plant Biotechnology ◽  
Plum Pox Virus ◽  
In Planta ◽  
Cross Hybridization ◽  
Promoter Sequences ◽  
Cp Gene ◽  
Virus Coat ◽  
Robust Response ◽  
Transgene Pyramiding

Our goal was to target silencing of the Plum pox virus coat protein (PPV CP) gene independently expressed in plants. Clone C-2 is a transgenic plum expressing CP. We introduced and verified, in planta, the effects of the inverse repeat of CP sequence split by a hairpin (IRSH) that was characterized in the HoneySweet plum. The IRSH construct was driven by two CaMV35S promoter sequences flanking the CP sequence and had been introduced into C1738 plum. To determine if this structure was enough to induce silencing, cross-hybridization was made with the C1738 clone and the CP expressing but PPV-susceptible C2 clone. In total, 4 out of 63 clones were silenced. While introduction of the IRSH is reduced due to the heterozygous character in C1738 plum, the silencing induced by the IRSH PPV CP is robust. Extensive studies, in greenhouse containment, demonstrated that the genetic resource of C1738 clone can silence the CP production. In addition, these were verified through the virus transgene pyramiding in the BO70146 BlueByrd cv. plum that successfully produced resistant BlueByrd BO70146 × C1738 (HybC1738) hybrid plums.

Transfer of Plum Pox Virus Coat Protein Genes from a Plum Pox-resistant Transgenic Clone of Prunus domestica Plum to Its Progeny through Hybridization

HortScience ◽  
1998 ◽  
Vol 33 (3) ◽  
pp. 532a-532
Author(s):  
Ralph Scorza ◽  
Laurene Levy ◽  
Vern Damsteegt ◽  
Ann Callahan ◽  
Kevin Webb ◽  
...  
Keyword(s):  
Coat Protein ◽  
Economic Losses ◽  
Plum Pox Virus ◽  
Post Inoculation ◽  
Cp Gene ◽  
Multiple Copies ◽  
Virus Coat ◽  
Aphid Inoculation ◽  
The Stability ◽  
Transgenic Clone

Sharka or plum pox virus (PPV) is a major disease of stone fruit and causes severe economic losses in Europe. There is little resistance to PPV in most Prunus species, thus genetic engineering represents a potentially useful approach to obtain resistant germplasm. Transgenic plums containing the PPV coat protein (CP) or the related papaya ringspot virus (PRV)-CP gene were produced through Agrobacterium tumefaciens-mediated transformation. These transgenic plum clones were then evaluated for resistance to PPV infection in the greenhouse by graft or aphid inoculation with PPV. While symptoms of PPV appeared in most transgenic clones, all plants of PPV-CP transgenic clone C5 were symptomless and ELISA and immunocapture-reverse transcriptase PCR negative for over three years following inoculation with two strains of PPV (Ravelonandro et al., Plant Dis. 81:1231-1235, 1997). Clone C5, which contains multiple copies of the PPV-CP gene, was hybridized with PRV-CP transgenic plants or untransformed plum cultivars. Progeny were obtained containing no transgenes, only the PPV-CP, only the PRV-CP, or both the PRV-CP and PPV-CP transgenes. Seedlings were inoculated with PPV. At 5 and 11 months post-inoculation, seedlings containing the PPV-CP genes from C5 were symptomless and ELISA negative. Seedlings containing only PRV-CP transgenes or non-transformed controls showed symptoms of PPV infection and were ELISA positive. These results indicate that the PPV-CP transgenes can be transferred to progeny through hybridization and that these genes can impart resistance to PPV in transgenic seedlings. The inheritance of the multicopy inserts of the PPV-CP and PRV-CP transgenes is being analyzed. The combined effects of both transgenes on resistance to PPV and the stability of PPV resistance in the progeny of the resistant C5 transgenic line are currently under evaluation.

scholarly journals Plum Pox Virus Coat Protein Gene Intron-hairpin-RNA (ihpRNA) Constructs Provide Resistance to Plum Pox Virus in Nicotiana benthamiana and Prunus domestica

2007 ◽  
Vol 132 (6) ◽  
pp. 850-858 ◽  
Author(s):  
Jean-Michel Hily ◽  
Michel Ravelonandro ◽  
Vern Damsteegt ◽  
Carole Bassett ◽  
Cesar Petri ◽  
...  
Keyword(s):  
Coat Protein ◽  
Gene Silencing ◽  
Nicotiana Benthamiana ◽  
High Capacity ◽  
Plum Pox Virus ◽  
Prunus Domestica ◽  
Hairpin Rna ◽  
Woody Perennial ◽  
Virus Coat ◽  
Interfering Rna

Constructs with self-complementary sequences separated by an intron produce “hairpin” RNA [intron-hairpin-RNA (ihpRNA)] structures that efficiently elicit posttranscriptional gene silencing (PTGS). In the current study, the authors use this technology to confer resistance to plum pox virus (PPV) in herbaceous and woody perennial plants by silencing the PPV–coat protein (CP) gene. The authors confirmed the high capacity of ihpRNA constructs for inducing RNA silencing in Nicotiana benthamiana Domin., as more than 75% of the transformants displayed PTGS as evaluated by specific small interfering RNA (siRNA) production. The authors demonstrated that ihpRNA constructs provided PPV resistance, and they found a correlation between the length of the PPV sequence introduced in the ihpRNA constructs and the frequency of transgenic-resistant plants. Plants transformed with the full-length sequence produced a higher percentage of resistant lines. The authors further demonstrated for the first time that ihpRNA technology is applicable to a woody perennial species. A transgenic plum (Prunus domestica L.) PPV-CP ihpRNA line showed gene silencing characteristics (hypermethylation of the transgene sequence and specific siRNA production) and resistance to PPV infection 16 months after inoculation.

EXPRESSION OF THE PLUM POX VIRUS COAT PROTEIN REGION IN ESCHERICHIA COLI

1989 ◽  
pp. 131-132
Author(s):  
D. Mattanovich ◽  
G. Himmler ◽  
M. Laimer ◽  
A. da Camara Machado ◽  
V. Hanzer ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Coat Protein ◽  
Plum Pox Virus ◽  
Coat Protein Region ◽  
Virus Coat Protein ◽  
Virus Coat

EXPRESSION OF THE PLUM POX VIRUS COAT PROTEIN GENE IN NICOTIANA CLEVELANDII

1990 ◽  
pp. 577-580 ◽  
Author(s):  
M. Laimer da Câmara Machado ◽  
A. da Câmara Machado ◽  
D. Mattanovich ◽  
F. Regner ◽  
V. Hanzer ◽  
...  
Keyword(s):  
Coat Protein ◽  
Coat Protein Gene ◽  
Protein Gene ◽  
Plum Pox Virus ◽  
Virus Coat Protein ◽  
Virus Coat ◽  
Nicotiana Clevelandii

scholarly journals Induction of Particle Polymorphism by Cucumber Necrosis Virus Coat Protein Mutants In Vivo

2007 ◽  
Vol 82 (3) ◽  
pp. 1547-1557 ◽  
Author(s):  
Kishore Kakani ◽  
Ron Reade ◽  
Umesh Katpally ◽  
Thomas Smith ◽  
D'Ann Rochon
Keyword(s):  
Coat Protein ◽  
Rna Binding ◽  
In Planta ◽  
Necrosis Virus ◽  
Protein Mutants ◽  
Cucumber Necrosis Virus ◽  
Virus Coat ◽  
R Domain

ABSTRACT The Cucumber necrosis virus (CNV) particle is a T=3 icosahedron consisting of 180 identical coat protein (CP) subunits. Plants infected with wild-type CNV accumulate a high number of T=3 particles, but other particle forms have not been observed. Particle polymorphism in several T=3 icosahedral viruses has been observed in vitro following the removal of an extended N-terminal region of the CP subunit. In the case of CNV, we have recently described the structure of T=1 particles that accumulate in planta during infection by a CNV mutant (R1+2) in which a large portion of the N-terminal RNA binding domain (R-domain) has been deleted. In this report we further describe properties of this mutant and other CP mutants that produce polymorphic particles. The T=1 particles produced by R1+2 mutants were found to encapsidate a 1.9-kb RNA species as well as smaller RNA species that are similar to previously described CNV defective interfering RNAs. Other R-domain mutants were found to encapsidate a range of specifically sized less-than-full-length CNV RNAs. Mutation of a conserved proline residue in the arm domain near its junction with the shell domain also influenced T=1 particle formation. The proportion of polymorphic particles increased when the mutation was incorporated into R-domain deletion mutants. Our results suggest that both the R-domain and the arm play important roles in the formation of T=3 particles. In addition, the encapsidation of specific CNV RNA species by individual mutants indicates that the R-domain plays a role in the nature of CNV RNA encapsidated in particles.

scholarly journals Transgenic or Plant Expression Vector-Mediated Recombination of Plum Pox Virus

2000 ◽  
Vol 74 (16) ◽  
pp. 7462-7469 ◽  
Author(s):  
Mark Varrelmann ◽  
Laszlo Palkovics ◽  
Edgar Maiss
Keyword(s):  
Potato Virus ◽  
Expression Vector ◽  
Wild Type Virus ◽  
Plum Pox Virus ◽  
Type Virus ◽  
Wild Type ◽  
Plant Line ◽  
Plant Expression ◽  
Plant Expression Vector ◽  
Cp Gene

ABSTRACT Different mutants of an infectious full-length clone (p35PPV-NAT) of Plum pox virus (PPV) were constructed: three mutants with mutations of the assembly motifs RQ and DF in the coat protein gene (CP) and two CP chimeras with exchanges in the CP core region ofZucchini yellow mosaic virus and Potato virus Y. The assembly mutants were restricted to single infected cells, whereas the PPV chimeras were able to produce systemic infections inNicotiana benthamiana plants. After passages in different transgenic N. benthamiana plants expressing the PPV CP gene with a complete (plant line 4.30.45.) or partially deleted 3′-nontranslated region (3′-NTR) (plant line 17.27.4.), characterization of the viral progeny of all mutants revealed restoration of wild-type virus by recombination with the transgenic CP RNA only in the presence of the complete 3′-NTR (4.30.45.). Reconstitution of wild-type virus was also observed following cobombardment of different assembly-defective p35PPV-NAT together with a movement-defective plant expression vector of Potato virus X expressing the intact PPV-NAT CP gene transiently in nontransgenic N. benthamiana plants. Finally, a chimeric recombinant virus was detected after cobombardment of defective p35PPV-NAT with a plant expression vector-derived CP gene from the sour cherry isolate of PPV (PPV-SoC). This chimeric virus has been established by a double recombination event between the CP-defective PPV mutant and the intact PPV-SoC CP gene. These results demonstrate that viral sequences can be tested for recombination events without the necessity for producing transgenic plants.

scholarly journals Resistance of Transgenic Prunus domestica to Plum Pox Virus Infection

Plant Disease ◽  
1997 ◽  
Vol 81 (11) ◽  
pp. 1231-1235 ◽  
Author(s):  
M. Ravelonandro ◽  
R. Scorza ◽  
J. C. Bachelier ◽  
G. Labonne ◽  
L. Levy ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Coat Protein ◽  
Plum Pox Virus ◽  
Prunus Domestica ◽  
Chain Reaction ◽  
Cp Gene ◽  
Aphid Feeding ◽  
Polymerase Chain ◽  
Dormancy Cycles ◽  
Das Elisa

Transgenic plum trees (Prunus domestica) containing the plum pox potyvirus coat protein (PPV-CP) gene were inoculated with PPV by aphid feeding or chip budding. Infection was monitored by evaluation of virus symptoms, DAS-ELISA, and immunoblot assays. Based on observations and analyses over 3 years including two dormancy cycles, one out of five transgenic clones (C-5), was found to be resistant to infection whether inoculated by aphids or by chip budding. PPV could not be detected in any inoculated plants of the C-5 clone by immunoblot or immunocap-ture-reverse transcriptase-polymerase chain reaction assays. To our knowledge, this is the first P. domestica clone resistant to PPV infection produced by genetic engineering.

Molecular cloning and characterization of Dbr1, a 2-alkenal reductase from Artemisia annuaThe nucleotide sequence reported in this article has been deposited in the GenBank database under accession No. FJ750460.This paper is one of a selection of papers published in a Special Issue from the National Research Council of Canada – Plant Biotechnology Institute.

Botany ◽  
2009 ◽  
Vol 87 (6) ◽  
pp. 643-649 ◽  
Author(s):  
Yansheng Zhang ◽  
Keat H. Teoh ◽  
Darwin W. Reed ◽  
Patrick S. Covello
Keyword(s):  
Double Bond ◽  
Artemisia Annua ◽  
Sequence Similarity ◽  
National Research Council ◽  
Plant Biotechnology ◽  
In Planta ◽  
Carbon Double Bond ◽  
Expressed Sequence ◽  
Selection Of

The molecular genetics of carbon–carbon double bond reduction in the plant Artemisia annua  L. was studied. Expressed sequence tags from this plant were investigated for sequences with similarity to known double-bond reductases. This resulted in the isolation of a cDNA, corresponding to the gene A. annua Dbr1 (Double bond reductase1), encoding a member of the medium chain dehydrogenase/reductase protein superfamily with sequence similarity to tobacco allyl alcohol dehydrogenase. Recombinant A. annua Dbr1 protein was purified from Escherischia coli and shown to catalyze the reduction of the carbon–carbon double bond of 2-alkenals. This activity included the reduction of the double bond at C11–C13 in the artemisinin precursor artemisinic aldehyde, albeit with unnatural stereochemistry. The substrate specificity, product stereochemistry, and expression pattern of A. annua Dbr1 point to its involvement in planta in the detoxification of 2-alkenals, which may be generated under oxidative stress conditions.

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