Construction, production, and purification of recombinant adenovirus vectors.
Methods Mol Biol
; 1089: 159-73, 2014.
Article
em En
| MEDLINE
| ID: mdl-24132485
ABSTRACT
Recombinant adenoviruses provide a versatile system for gene expression studies and therapeutic applications. In this chapter, a standard procedure for their generation and small-scale production is described. Homologous recombination in E. coli between shuttle plasmids and full-length adenovirus backbones (E1-deleted) is used for the generation of recombinant adenoviral vectors genomes. The adenovirus genomes are then analyzed to confirm their identity and integrity, and further linearized and transfected to generate a recombinant adenoviral vector in permissive human cells. These vectors are then purified by two sequential CsCl gradient centrifugations and subjected to a chromatography step in order to eliminate the CsCl and exchange buffers. Finally, the viral stock is characterized through the quantification of its viral particle content and its infectivity.
Texto completo:
1
Coleções:
01-internacional
Temas:
Geral
Base de dados:
MEDLINE
Assunto principal:
Adenovírus Humanos
/
Vetores Genéticos
Limite:
Humans
Idioma:
En
Revista:
Methods Mol Biol
Assunto da revista:
BIOLOGIA MOLECULAR
Ano de publicação:
2014
Tipo de documento:
Article
País de afiliação:
Espanha