A selectable and excisable marker system for the rapid creation of recombinant poxviruses.
PLoS One
; 6(9): e24643, 2011.
Article
em En
| MEDLINE
| ID: mdl-21931792
ABSTRACT
BACKGROUND:
Genetic manipulation of poxvirus genomes through attenuation, or insertion of therapeutic genes has led to a number of vector candidates for the treatment of a variety of human diseases. The development of recombinant poxviruses often involves the genomic insertion of a selectable marker for purification and selection purposes. The use of marker genes however inevitably results in a vector that contains unwanted genetic information of no therapeutic value. METHODOLOGY/PRINCIPALFINDINGS:
Here we describe an improved strategy that allows for the creation of marker-free recombinant poxviruses of any species. The Selectable and Excisable Marker (SEM) system incorporates a unique fusion marker gene for the efficient selection of poxvirus recombinants and the Cre/loxP system to facilitate the subsequent removal of the marker. We have defined and characterized this new methodological tool by insertion of a foreign gene into vaccinia virus, with the subsequent removal of the selectable marker. We then analyzed the importance of loxP orientation during Cre recombination, and show that the SEM system can be used to introduce site-specific deletions or inversions into the viral genome. Finally, we demonstrate that the SEM strategy is amenable to other poxviruses, as demonstrated here with the creation of an ectromelia virus recombinant lacking the EVM002 gene. CONCLUSION/SIGNIFICANCE:
The system described here thus provides a faster, simpler and more efficient means to create clinic-ready recombinant poxviruses for therapeutic gene therapy applications.
Texto completo:
1
Bases de dados:
MEDLINE
Assunto principal:
Poxviridae
/
Recombinação Genética
/
Marcadores Genéticos
Limite:
Animals
/
Humans
Idioma:
En
Revista:
PLoS One
Assunto da revista:
CIENCIA
/
MEDICINA
Ano de publicação:
2011
Tipo de documento:
Article
País de afiliação:
Canadá