Construction, identification and expression of recombinant eukaryotic vector pEGFP-BMI-1 / 中国实验血液学杂志
Journal of Experimental Hematology
;
(6): 1056-1060, 2007.
Artigo
em Chinês
| WPRIM
| ID: wpr-318790
ABSTRACT
This study was purposed to construct and identify the mammalian expression vector of pEGFP-BMI-1 and to detect whether it could express in human cervix cancer cell line HeLa. The cDNA fragment of BMI-1 obtained by RT-PCR was inserted into pEGFP-N1. The recombinant plasmid was confirmed by restriction enzyme digestion, PCR and DNA sequencing. pEGFP-BMI-1 was transfected into HeLa cells with lipofectamine 2000. The expression of pEGFP-BMI-1 was determined by EGFP fluorescence and Western blot analysis. SYBR Green I real-time RT-PCR was used to quantitate P16INK4a mRNA. The results showed that the correct construction of the recombinant plasmid pEGFP-BMI-1 has been shown by restriction enzyme digestion, PCR and DNA sequencing. pEGFP-BMI-1 could express BMI-1-EGFP fusion protein in HeLa cells. Real-time RT-PCR showed that P16INK4a mRNA expression was reduced to 9.2%. It is concluded that the vector of pEGFP-BMI-1 has been successfully constructed and it can be expressed in HeLa cells. This work has laid foundations for further study on biological functions and potential application of BMI-1.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Proteínas Repressoras
/
Proteínas Recombinantes de Fusão
/
RNA Mensageiro
/
Dados de Sequência Molecular
/
Proteínas Nucleares
/
Sequência de Bases
/
Células HeLa
/
Transfecção
/
Proteínas Proto-Oncogênicas
/
Inibidor p16 de Quinase Dependente de Ciclina
Tipo de estudo:
Estudo prognóstico
Limite:
Humanos
Idioma:
Chinês
Revista:
Journal of Experimental Hematology
Ano de publicação:
2007
Tipo de documento:
Artigo
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