[Construction of fusion expression vector of human-derived neurotrophin-6 gene encoding mature peptide and purification of its expressed product].
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.
Zhang
Chengwu
C
Department of Physiology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Cai
Qingsong
Q
Li
Zicheng
Z
Zhai
Chaoyang
C
Zheng
Yu
Y
D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D009414:Nerve Growth Factors / Q000096:biosynthesis* / Q000235:genetics; D010455:Peptides / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*
D009414:Nerve Growth Factors; D010455:Peptides; D011993:Recombinant Fusion Proteins; C075936:neurotrophin 6; D007544:Isopropyl Thiogalactoside