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Establishment of a Plasmid-Based Reverse Genetics System for the Cell Culture-Adapted Hepatitis E Virus Genotype 3c Strain 47832c
- Source :
- Pathogens, Vol 9, Iss 3, p 157 (2020), Pathogens, Volume 9, Issue 3
- Publication Year :
- 2020
- Publisher :
- MDPI AG, 2020.
-
Abstract
- The hepatitis E virus (HEV) causes acute and chronic hepatitis in humans. Investigation of HEV replication is hampered by the lack of broadly applicable, efficient cell culture systems and tools for site-directed mutagenesis of HEV. The cell culture-adapted genotype 3c strain 47832c, which represents a typical genotype predominantly detected in Europe, has previously been used for several basic and applied research studies. Here, a plasmid-based reverse genetics system was developed for this strain, which efficiently rescued the infectious virus without the need for in vitro RNA transcription. The cotransfection of T7 RNA polymerase-expressing BSR/T7 cells with one plasmid encoding the full-length viral genome and two helper plasmids encoding vaccinia virus capping enzymes resulted in the production of infectious HEV, which could be serially passaged on A549/D3 cells. The parental and recombinant virus exhibited similar replication kinetics. A single point mutation creating an additional restriction enzyme site could be successfully introduced into the virus genome of progeny virus, indicating that the system is suitable for site-directed mutagenesis. This system is the first plasmid-based HEV reverse genetics system, as well as the first reverse genetics system for HEV genotype 3c, and should therefore be of broad use for basic and applied HEV research.
- Subjects :
- Microbiology (medical)
reverse genetics system
viruses
Mutagenesis (molecular biology technique)
lcsh:Medicine
Biology
medicine.disease_cause
Recombinant virus
Virus
Article
chemistry.chemical_compound
Plasmid
hepatitis e virus
Hepatitis E virus
Genotype
medicine
Immunology and Allergy
Molecular Biology
cell culture
General Immunology and Microbiology
lcsh:R
Virology
genotype 3c
Reverse genetics
Infectious Diseases
chemistry
Vaccinia
site-directed mutagenesis
Subjects
Details
- Language :
- English
- ISSN :
- 20760817
- Volume :
- 9
- Issue :
- 3
- Database :
- OpenAIRE
- Journal :
- Pathogens
- Accession number :
- edsair.doi.dedup.....3e6fc8bb05fb96a1e16db40f4e67f1e3