Expression, purification and polyclonal antibody preparation for a novel gene BC097361.
- Author:
Qi WANG
1
;
Yun-lei LIANG
;
Hong-shan WEI
;
Hui-chun XING
;
Jun CHENG
;
Meng-dong LAN
;
Bin ZHANG
Author Information
- Publication Type:Journal Article
- MeSH: Angiotensin II; genetics; Animals; Antibodies; immunology; isolation & purification; metabolism; Antibody Specificity; Blotting, Western; Cloning, Molecular; Escherichia coli; genetics; metabolism; Gene Expression; Genetic Vectors; genetics; Liver Cirrhosis; genetics; Male; Plasmids; genetics; Rabbits; Recombinant Fusion Proteins; biosynthesis; immunology; Reverse Transcriptase Polymerase Chain Reaction
- From: Chinese Journal of Hepatology 2009;17(8):589-593
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo express and purify of the BC097361 recombinant protein, and to prepare the BC097361 specific rabbit polyclonal antibody.
METHODSBC097361 cDNA was ligated into the prokaryotic expressive vector pET-32a (+), and the resulting plasmid was transformed into E.coli BL21 (DE3). The protein expression was induced with IPTG and the protein was analyzed with SDS-PAGE and western blotting. The expressed product was purified using Ni+ affinity column chromatography.Then the purified pET-32a (+) -BC097361 fusion protein was used to immunize New Zealand rabbits to gain polyclonal antibody. The specificity and potency of polyclonal antibody were evaluated by Western blot and ELISA.
RESULTSThe BC097361 fusion protein was highly expressed.The protein was mainly in the inclusion body. ELISA indicated the titer of polyclonal antibody more than 1:320000. The high specificity was confirmed with Western blot.
CONCLUSIONSThe recombinant BC097361 fusion protein and the BC097361 specific polyclonal antibody will be valuable tools for the investigation on the biological function of BC097361.
