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Production of Lentiviral Vectors for Transducing Cells from the Central Nervous System
Published on: May 24, 2012
Essential and dispensable virus-encoded replication elements revealed by efforts To develop hypoviruses as gene
N Suzuki1, L M Geletka, D L Nuss
1Center for Agricultural Biotechnology, University of Maryland Biotechnology Institute, University of Maryland, College Park, Maryland 20742, USA.
Journal of Virology
|July 25, 2000
Summary
Hypoviruses, used to attenuate chestnut blight fungus, were tested as gene expression vectors. While transient expression occurred, long-term gene expression was unsuccessful due to foreign gene deletion, revealing insights into hypovirus replication.
Area of Science:
- Mycology
- Virology
- Molecular Biology
Background:
- Hypoviruses cause virulence attenuation in the chestnut blight fungus Cryphonectria parasitica.
- Investigating hypoviruses as potential gene expression vectors is a novel approach.
Purpose of the Study:
- To assess the feasibility of using hypoviruses as gene expression vectors.
- To identify essential and dispensable elements for hypovirus replication.
Main Methods:
- Modification of the CHV1-EP713 hypovirus cDNA clone to create vector candidates.
- Transfection of fungal colonies and analysis of viral RNA for foreign gene expression and retention.
- Characterization of vector viability and progeny to understand replication determinants.
Main Results:
- Transient expression of foreign genes (e.g., GFP) was achieved but not sustained long-term.
- Foreign genes were readily deleted from replicating hypoviruses, though some sequence remnants persisted.
- The N-terminal 24 codons of ORF A are essential for replication, while the rest is dispensable.
- Hypovirus replication tolerated significant alterations in ORF A termination and ORF B initiation regions.
Conclusions:
- Hypoviruses are not currently suitable as long-term gene expression vectors due to efficient foreign sequence deletion.
- Essential regions for hypovirus replication and sequence deletion mechanisms were identified.
- Findings provide insights into hypovirus biology and potential vector development.
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