Construction and selection of effective mouse Smad6 recombinant lenti-virus interference vectors / 生物医学工程学杂志
Journal of Biomedical Engineering
; (6): 1100-1104, 2010.
Article
en Zh
| WPRIM
| ID: wpr-260930
Biblioteca responsable:
WPRO
ABSTRACT
This experiment was designed to construct mouse Smad6 recombinant RNA interference vectors and determine their interference effects on bone marrow mesenchymal stem cells (BMSCs). Three recombinant Smad6 RNA interference vectors were constructed by molecular clone techniques with a lenti-virus vector expressing green fluorescent protein (GFP), and the correctness of recombinant vectors was verified by DNA sequencing. Mouse BMSCs were used for transfection experiments and BMP-2 was in use for osteogenic induction of MSCs. The transfection efficiency of recombinant vectors was examined by Laser confocal scanning microscope and the interference effect of recombinant vectors on Smad6 gene expression was determined by real-time RT-PCR and Western blot, respectively. Three Smad6 recombinant RNA interference vectors were successfully constructed and their correctness was proved by DNA sequencing. After transfection, GFPs were effectively expressed in MSCs and all of three recombinant vectors gained high transfection efficiency (> 95%). Both real-time PCR and Western blot examination indicated that among three recombinant vectors, No. 2 Svector had the best interference effect and the interference effect was nearly 91% at protein level. In conclusion, Mouse recombinant Smad6 RNA interference (RNAi) vector was successfully constructed and it provided an effective tool for further studies on BMP signal pathways.
Texto completo:
1
Índice:
WPRIM
Asunto principal:
Proteínas Recombinantes
/
Transfección
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Lentivirus
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Proteínas Morfogenéticas Óseas
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Interferencia de ARN
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Proteínas Fluorescentes Verdes
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Proteína smad6
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Células Madre Mesenquimatosas
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Vectores Genéticos
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Genética
Límite:
Animals
Idioma:
Zh
Revista:
Journal of Biomedical Engineering
Año:
2010
Tipo del documento:
Article