In Vitro Amplification and Selection of Engineered RNase P Ribozyme for Gene Targeting Applications.
Methods Mol Biol
; 2822: 419-429, 2024.
Article
en En
| MEDLINE
| ID: mdl-38907932
ABSTRACT
Ribozymes engineered from the RNase P catalytic RNA (M1 RNA) represent promising gene-targeting agents for clinical applications. We describe in this report an in vitro amplification and selection procedure for generating active RNase P ribozyme variants with improved catalytic efficiency. Using the amplification and selection procedure, we have previously generated ribozyme variants that were highly active in cleaving a herpes simplex virus 1-encoded mRNA in vitro and inhibiting its expression in virally infected human cells. In this chapter, we use an overlapping region of the mRNAs for the IE1 and IE2 proteins of human cytomegalovirus (HCMV) as a target substrate. We provide detailed protocols and include methods for establishing the procedure for the amplification and selection of active mRNA-cleaving RNase P ribozymes. The in vitro amplification and selection system represents an excellent approach for engineering highly active RNase P ribozymes that can be used in both basic research and clinical applications.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
ARN Catalítico
/
Marcación de Gen
/
Ribonucleasa P
Límite:
Humans
Idioma:
En
Revista:
Methods Mol Biol
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2024
Tipo del documento:
Article
País de afiliación:
Estados Unidos
Pais de publicación:
Estados Unidos