[Construction of pcDNA3.1(+)/A2E eukaryotic expression vector and its expression on K562 cell].
Zhongguo Shi Yan Xue Ye Xue Za Zhi
; 13(3): 464-7, 2005 Jun.
Article
em Zh
| MEDLINE
| ID: mdl-15972143
ABSTRACT
To construct pcDNA3.1(+)/A2E eukaryotic expression vector and obtain a stable expression on HLA-I negative human K562 cell, PCR technique was employed to amplify A2E cDNA from the multi-cistron expression vector pG/A2E carrying HLA-E and HLA-A2 cDNA through internal ribozyme entry site (IRES), the cDNA was subcloned into vector pcDNA3.1(+), thus a eukaryotic expression was constructed and named pcDNA3.1(+)/A2E; then, the recombinant plasmid was transferred into the target cells, followed by screening with G418 and limiting dilution; finally, flow cytometry was adopted to detect HLA-E expression on the target cells. The results showed that HLA-E molecules were successfully expressed on K562 cells transfected with pcDNA3.1(+)/A2E (27.76%) and the expression of HLA-E molecules was not detected on K562 cells transfected with pcDNA3.1(+). It is concluded that the pcDNA 3.1(+)/A2E eukaryotic expression vector was successfully constructed and the HLA-E molecules were expressed on K562 cells. The data presented here would be expected to lay a good basis for the research of the molecular mechanism of HLA-E function and the interaction between HLA-E and the receptor on NK cells, as well as the influence of the expression of HLA-E in vitro on NK cells.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Antígenos de Histocompatibilidade Classe I
/
Vetores Genéticos
/
Antígenos HLA
Limite:
Humans
Idioma:
Zh
Revista:
Zhongguo Shi Yan Xue Ye Xue Za Zhi
Ano de publicação:
2005
Tipo de documento:
Article