Construction of nonsense-mutated eukaryotic expression vector of factor IX gene and its expression in COS-7 cells / 中国实验血液学杂志
Journal of Experimental Hematology
;
(6): 749-752, 2010.
Artículo
en Chino
| WPRIM
| ID: wpr-237659
ABSTRACT
The purpose of this study was to construct 4 types of nonsense-mutated eukaryotic expression plasmids of fIX gene, using pcDNA3.1 plasmid containing fIX cDNA as template, and to identify, then to perform their expression in COS-7 cells. These stop mutants constructed by site-directed mutagenesis based on PCR, and further confirmed by DNA sequencing. COS-7 cells were transfected with either the wild-type or mutated fIX expression constructs, then the relative expression levels of fIX mRNA were detected by real time fluorescent quantitative PCR. The result showed that except the designed sites, there were no other nucleotide mutation in the sequences of four nonsense mutants. The results of real time PCR proved that the nonsense-mutated vectors can be effectively expressed in COS-7 cells. It is concluded that the nonsense-mutated eukaryotic expression vectors of fIX gene have been successfully constructed and can express in COS-7 cells, which provides the material basis for further researches on mechanism and treatment of FIX deficiency and the function defects caused by nonsense mutation.
Texto completo:
Disponible
Índice:
WPRIM (Pacífico Occidental)
Asunto principal:
Factor IX
/
Secuencia de Bases
/
Transfección
/
Chlorocebus aethiops
/
Clonación Molecular
/
Codón sin Sentido
/
Células COS
/
Vectores Genéticos
/
Genética
Tipo de estudio:
Estudio pronóstico
Límite:
Animales
Idioma:
Chino
Revista:
Journal of Experimental Hematology
Año:
2010
Tipo del documento:
Artículo
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