Construction of pGEX4T-1-Cox7a2 and expression, purification and identification of the recombinant protein.
- Author:
Liang CHEN
1
;
Zhong-cheng XIN
;
Xue-jun JIANG
;
Long TIAN
;
Yi-ming YUAN
;
Gang LIU
;
Wei-dong SONG
;
Ying-lu GUO
Author Information
- Publication Type:Journal Article
- MeSH: Animals; Cell Line; Cloning, Molecular; Electron Transport Complex IV; biosynthesis; genetics; isolation & purification; Genetic Vectors; Leydig Cells; metabolism; Male; Mice; Mitochondria; physiology; Recombinant Fusion Proteins; biosynthesis; isolation & purification; Reverse Transcriptase Polymerase Chain Reaction
- From: National Journal of Andrology 2006;12(9):794-797
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo clone and express Cox7a2, one mitochondrial respiratory chain related gene, and to identify its recombinant protein.
METHODSThe coding region of Cox7a2 was amplified from primary cultured mouse Leydig cells by RT-PCR. The PCR product was cloned into pGEX4T-1 vector by BamH I and EcoR I sites, and confirmed by DNA sequencing. The recombinant fusion protein vector was transformed and expressed into BL21. The recombinant fusion protein was identified by Western blotting.
RESULTSThe entire coding region of Cox7a2 was cloned and expressed. The fusion protein was identified by anti-GST monoclonal antibody using Western blotting.
CONCLUSIONThe cloning of Cox7a2 and the expression of the recombinant protein would help to study the detailed function of Cox7a2, one respiratory chain related and highly differently expressed gene in the tissues of aging testes.
