Construction and expression of fusion protein TRX-hJagged1 in E.coli BL21.
10.7534/j.issn.1009-2137.2014.03.043
- Author:
Guo-Hui LI
1
;
Yu-Zhen FAN
2
;
Si-Yong HUANG
1
;
Qiang LIU
1
;
Dan-Dan YIN
1
;
Li LIU
1
;
Ren-An CHEN
1
;
Miao-Wang HAO
1
;
Ying-Min LIANG
3
Author Information
1. Department of Hematology, Tangdu Hospital of The Fourth Military Medicol University, Xian 710038, Shaanxi.
2. Department of Traditional Chinese Medicine, Tangdu Hospital of The Fourth Military Medicol University, Xian 710038, Shaanxi.
3. Department of Hematology, Tangdu Hospital of The Fourth Military Medicol University, Xian 710038, Shaanxi. E-mail: liangym@fmmu.edu.cn.
- Publication Type:Journal Article
- MeSH:
Calcium-Binding Proteins;
genetics;
metabolism;
Cloning, Molecular;
Escherichia coli;
metabolism;
Genetic Vectors;
Intercellular Signaling Peptides and Proteins;
genetics;
metabolism;
Jagged-1 Protein;
Membrane Proteins;
genetics;
metabolism;
Plasmids;
Recombinant Fusion Proteins;
biosynthesis;
genetics;
Serrate-Jagged Proteins
- From:
Journal of Experimental Hematology
2014;22(3):807-811
- CountryChina
- Language:Chinese
-
Abstract:
This study was purposed to construct prokaryotic expression vector and to investigate the expression of Notch ligand Jagged1 in E.coli. An expression vector pET-hJagged1 was constructed, which can be inserted in Jagged1 with different lengths, but the DSL domain of human Jagged1 should be contained. Then the recombinant plasmids were transformed into the competent cell of E.coli BL21, and the expression of the fusion protein was induced by IPTG. Fusion protein was purified from the supernatant of cell lysates via the Nickel affinity chromatography. The results showed that prokaryotic expression vectors pET-hJagged1 (Bgl II), pET-hJagged1 (Hind I) and pET-hJagged1 (Stu I) were successfully constructed, but only pET-hJagged1 (Stu I) could express the soluble TRX-hJagged1. The purified TRX-Jagged1 protein could be obtained via the Nickel affinity chromatography, and then confirmed by Western Blot. It is concluded that prokaryotic expression vector pET-hJagged1 is successfully constructed, but only pET-hJagged1 (Stu I) can express the soluble TRX-hJagged1 and the TRX-Jagged1 fusion protein is obtained through the prokaryotic expression system, which laid a solid foundation for further to explore the effects of Jagged1 in hematopoietic and lymphoid system.
