[The expression of fusion protein GST-GP302 in E.coli and its preparation of rabbit anti-serum].
OBJECTIVE
To express fusion protein of GST and vWf binding domain(GP302) of platelet GPIbalpha in E.coli and its preparation of rabbit anti-serum.


METHODS
GP302 gene was inserted into pGEX-4T-1. The recombinant vector was identificated by restriction endonuclease digestion analysis. Fusion protein GST-GP302 was expressed in E.coli via IPTG induction. The rabbit antibody against GST-GP302 was prepared by using renatured GST-GP302 as immuneogen and the specificity of polyclonal antibody was identified by Western blot.


RESULTS
The restriction endonuclease digestion analysis of recombinant plasmid demonstrated that the GP302 gene had been exactly inserted into pGEX-4T-1. SDS-PAGE analysis showed that the ralative molecular mass(M(r)) of the fusion protein was about 59 000. ELISA analysis proved that the titer of rabbit serum against GST-GP302 was 10(-5). The polyclonal antibody specifically bound to purified platelet GPIbalpha.


CONCLUSIONS
The preparation of polyclonal antibody against GP302 peptide provides an usefal reagent for the detection of platelet GPIbalpha.
Su
Li
L
Institute of Transfusion, Academy of Military Medical Sciences, Beijing 100850, China.
Zhao
Jing-Xiang
JX
Wang
Bo
B
Zhang
Yu-Hua
YH
Guan
Li-Dong
LD
Wang
Zi-Ling
ZL
D000818:Animals; D000906:Antibodies / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics
D000906:Antibodies; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins