Engineering of alfalfa mosaic virus RNA 3 into an expression vector
| Autor: | A. Ragozzino, R. Miglino, Jesús A. Sánchez-Navarro, John F. Bol |
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| Přispěvatelé: | SANCHEZ NAVARRO, J., Miglino, R., Ragozzino, Antonio, Bol, J. F. |
| Rok vydání: | 2001 |
| Předmět: |
DNA
Complementary Recombinant Fusion Proteins Genetic Vectors Green Fluorescent Proteins Meristem Molecular Sequence Data Biology Viral Proteins Capsid Genes Reporter Virology Alfalfa mosaic virus Sequence Homology Nucleic Acid Gene expression Tobacco Genes Synthetic Gene Silencing Movement protein Promoter Regions Genetic Gene 3' Untranslated Regions Polymerase Subgenomic mRNA Plant Diseases Expression vector Base Sequence Virulence Protoplasts fungi RNA General Medicine biology.organism_classification Plants Genetically Modified Protein Structure Tertiary Plant Leaves Plant Viral Movement Proteins Luminescent Proteins Plants Toxic biology.protein Nucleic Acid Conformation RNA Viral Capsid Proteins Genetic Engineering |
| Zdroj: | Archives of virology. 146(5) |
| ISSN: | 0304-8608 |
| Popis: | RNA 3 of alfalfa mosaic virus (AMV) encodes the 5′-proximal movement protein (MP) gene and the 3′-proximal coat protein (CP) gene which is expressed from a subgenomic RNA. Several strategies were explored to use this RNA as a vector for expression of the green fluorescent protein (GFP) in Nicotiana tabaccum plants expressing the viral polymerase proteins P1 and P2 (P12 plants). Insertion of a subgenomic promoter (sgp)-GFP cassette between the CP gene and the 3′-untranslated region (UTR) interfered with RNA accumulation in protoplasts, indicating that cis-acting sequences required for replication were disrupted. When GFP was fused to the N-terminus of MP or CP, the chimeric RNAs accumulated in protoplasts but cell-to-cell movement in plants was blocked. Insertion of a GFP-sgp cassette immediately upstream of the CP gene caused a hypersensitive host response. However, insertion of a GFP-sgp cassette upstream of the MP gene did not affect symptom formation and yielded a vector that expressed GFP in inoculated but not in the systemic leaves of both P12 tobacco and non-transgenic N. benthamina plants. When the size of the GFP gene was reduced from 700 to 300 nucleotides, virus infection was observed in the non-inoculated leaves. Analysis of the progeny of some chimera revealed novel data on replication, encapsidation and recombination of AMV RNA 3. |
| Databáze: | OpenAIRE |
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