Construction of fusion expression vector of human-derived neurotrophin-6 gene encoding mature peptide and purification of its expressed product.
- Author:
Chengwu ZHANG
1
;
Qingsong CAI
;
Zicheng LI
;
Chaoyang ZHAI
;
Yu ZHENG
Author Information
1. Department of Physiology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
- Publication Type:Journal Article
- MeSH:
Escherichia coli;
genetics;
metabolism;
Genetic Vectors;
genetics;
Humans;
Isopropyl Thiogalactoside;
pharmacology;
Nerve Growth Factors;
biosynthesis;
genetics;
Peptides;
genetics;
Recombinant Fusion Proteins;
biosynthesis;
genetics;
isolation & purification
- From:
Journal of Biomedical Engineering
2005;22(6):1241-1244
- CountryChina
- Language:Chinese
-
Abstract:
To get the mature peptide of human-derived neurotrophin-6 (NT-6), NT-6 gene encoding mature peptide was amplified by PCR, using the NT-6 cDNA that had been cloned as templet. The gene encoding mature peptide of NT-6 gene was cloned into pGEX1-lambdaT plasmid to construct the fusion expression vector. Expression of fusion protein in Escherichia coli was defected after induction by isopropyl beta-D-thiogalactoside(IPTG). The mature peptide of NT-6 was collected with GST fusion protein purifying kit. It was shown that a fragment of 460bp was gained by PCR. With the techniques of double-cleave and electrophoresis, the recombinant vector was identified as pGEX1-NT-6. The recombinant vector pGEX1-NT-6 transformed Escherichia coli expressed fusion protein of 41KD after induction by IPTG. Cleaved by thrombin, the mature peptide of NT-6 was obtained; its molecular weight was about 15KD. The cloning and expression of human-derived NT-6 gene encoding mature protein has provided a basis for further studies on the function and clinical application of NT-6.
