Preparation of recombinant protein of human vascular endothelial cell growth inhibition factor rhVEGI-192A

Abstract

The invention provides a method for preparing recombinant protein of a human vascular endothelial cell-growth inhibiting factor rhVEGI-192A. The method comprises the steps of using a nucleotide sequence of VEGI-192A gene coding to construct an expression vector and inducing the expression vector to express in a host cell. Particularly, total RNA of human umbilical vein vascular endothelial cell line HUVEC is amplified to obtain a VEGI-192A gene fragment; a target gene is obtained through digestion by use of restriction endonuclease double enzyme; the target gene is connected to the expressionvector pET-30a to construct a recombinant protein expression vector; and the recombinant protein expression vector is transformed into escherichia coli host bacteria to induce expression so as to obtain purified protein. The method has the advantages of building a target-protein prokaryotic expression system with high efficiency, high yield, high activity and high purity on the basis of maintaining the natural spatial conformation of rhVEGI-192 recombinant protein, optimizing expression conditions and building a method for separating, purifying and renaturing target products.

Description

technical field

[0001] The invention relates to the field of biotechnology, in particular to a method for preparing a recombinant protein of human vascular endothelial cell growth inhibitory factor rhVEGI-192. Background technique

[0002] Vascular endothelial cell growth inhibitory factor (VEGI) is a recently discovered angiogenesis inhibitory factor, mainly produced by vascular endothelial cells. In 1997, Tan et al. first discovered human VEGI by screening the cDNA library of human umbilical vein endothelial cells, and named it TLI (TNF-like ligand 1) at that time. Subsequent biological activity research found that VEGI can significantly inhibit the proliferation of endothelial cells, hence the name. The full-length VEGI gene is 17Kb, consisting of four exons I, II, III, and IV and three introns. Three kinds of mRNA were spliced ​​according to different splicing patterns, encoding VEGI-251, VEGI-192 and VEGI-174 isoforms composed of 251, 192 and 174 amino acid residues, ...

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