[Construction of pGEX4T-1-Cox7a2 and expression, purification and identification of the recombinant protein].
OBJECTIVE
To clone and express Cox7a2, one mitochondrial respiratory chain related gene, and to identify its recombinant protein.


METHODS
The 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.


RESULTS
The entire coding region of Cox7a2 was cloned and expressed. The fusion protein was identified by anti-GST monoclonal antibody using Western blotting.


CONCLUSIONS
The 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.
Chen
Liang
L
Andrology Center, the First Hospital of Peking University, Beijing 100009, China. bdyychenliang@163.com
Xin
Zhong-cheng
ZC
Jiang
Xue-jun
XJ
Tian
Long
L
Yuan
Yi-ming
YM
Liu
Gang
G
Song
Wei-dong
WD
Guo
Ying-lu
YL
D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular; D003576:Electron Transport Complex IV / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D005822:Genetic Vectors; D007985:Leydig Cells / Q000378:metabolism*; D008297:Male; D051379:Mice; D008928:Mitochondria / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction
D011993:Recombinant Fusion Proteins; C514296:Cox7a2 protein, mouse; D003576:Electron Transport Complex IV