Construction of eukaryotic recombinant vector and expression in COS7 cell of LipL32-HlyX fusion gene from Leptospira serovar Lai / 生物医学工程学杂志
Journal of Biomedical Engineering
;
(6): 385-389, 2009.
Artigo
em Chinês
| WPRIM
| ID: wpr-280194
ABSTRACT
This study was conducted to construct eukaryotic recombinant vector of LipL32-HlyX fusion gene from Leptospira serovar Lai and express it in mammalian cell. Both of LipL32 gene and HlyX gene were amplified from Leptospira strain O17 genomic DNA by PCR. Then with the two genes as template, LipL32-HlyX fusion gene was obtained by SOE PCR (gene splicing by overlap extension PCR). The fusion gene was then cloned into pcDNA3.1 by restriction nuclease digestion. Having been transformed into E. coli DH5alpha, the recombiant plasmid was identified by restriction nuclease digestion, PCR analysis and sequencing. The recombinant plasmid was then transfected into COS7 cell whose expression was detected by RT-PCR and Western blotting analysis. RT-PCR amplified a fragment about 2000 bp and Western blotting analysis found a specific band about 75 KD which was consistent with the expected fusion protein size. In conclusion, the successful construction of eukaryotic recombinant vector containing LipL32-HlyX fusion gene and the effective expression in mammalian have laid a foundation for the application of Leptospira DNA vaccine.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Proteínas da Membrana Bacteriana Externa
/
Proteínas Recombinantes de Fusão
/
Chlorocebus aethiops
/
Sequências Hélice-Alça-Hélice
/
Células COS
/
Fusão Gênica
/
Vetores Genéticos
/
Genética
/
Leptospira
/
Lipoproteínas
Limite:
Animais
Idioma:
Chinês
Revista:
Journal of Biomedical Engineering
Ano de publicação:
2009
Tipo de documento:
Artigo
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