Construction expression and purification of C reactive protein
- VernacularTitle:C反应蛋白质粒的构建表达及纯化
- Author:
Zhi-Nong YIN
;
Xin ZHOU
;
Fang ZHENG
;
Lei ZHAO
;
Yong XIA
;
Han-Ning HU
;
Xin-Qiang JIANG
;
- Publication Type:Journal Article
- Keywords:
C-Reactive protein;
Gene expression;
Western blot
- From:
Chinese Journal of Laboratory Medicine
2003;0(09):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To build a foundation for determination of C reaction protein,C reaction protein was expressed and purified,and the immune reactivity of the purified protein was identified.Methods The CRP cDNA was amplified by RT-PCR from human liver cDNA library and inserted into expression vector pCRTT/NT.The recombined plasmid CRP-pCRTT/NT which expressed the fusion protein of CRP was then transferred into lysogenic host strain E coli.BL21 (DE3).The target protein was identified using SDS- polyacrylamide gel electrophoresis (SDS-PAGE).Affinity chromatography was used for protein purification.The immune reactivity of purified CRP was identified by Western blot using anti-CRP specific antibody.Results Recombiant human CRP was expressed in inclusion bodies of E.coli with a six histamine tag.The purify of recombinant protein was detected by SDS-PAGE as a single band at 30 000 and was identified by Western blot.Conclusions A plasmid expressed CRP protein is constructed and the purification system of CRP protein is established.The immune reactivity of the purified protein is identified by Western blot,which makes a good base for the preparation of CRP test kit.
