Abstracts
Copyright ©The Author(s) 2000. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Sep 15, 2000; 6(Suppl3): 117-117
Published online Sep 15, 2000. doi: 10.3748/wjg.v6.iSuppl3.117
Construction, expression and characterization of double-copy genes of truncated form of human insulin-like growth factor-I
He-Ying Sun, Xiao-Hui Liu, Shu-Wen Liang
He-Ying Sun, Xiao-Hui Liu, Shu-Wen Liang, Department of Life Science, Shanxi University, Taiyuan 030006, Shanxi Province, China
Author contributions: All authors contributed equally to the work.
Supported by the grants from Science Foundation of Shanxi No. 951029 and Reback Science Foundation No. 94124
Correspondence to: Xiao-Hui Liu, Department of Life Science, Shanxi University, Taiyuan 030006, Shanxi Province, China. xhliu@sxu.edu.cn
Telephone: 351-7011562
Received: February 13, 2000
Revised: April 5, 2000
Accepted: May 10, 2000
Published online: September 15, 2000
Abstract

AIM: To increase the production of recombinant des (1-3) IGF-I by increasing the copy number of gene carried on an expression vector, and to partially purify the expressed des (1-3) IGF-I, as well as compare its bio-activity with standard IGF-I.

METHODS: Second copy of des (1-3) IGF-I gene was inserted into pExSec1/IGF-I expression vector constructed by our previous work and carryed already one des (1-3) IGF-I gene, to form PExSec1/2 (IGF-I) expression plasmid, which carried two copies of tandem des (1-3) IGF-I gene. This plasmid was transformed into a protease deficient E. coli strain BL21 (DE3). The engineered bacteria was cultured and induced at low temperature. The expressed product was purified through ultra-filtration and gel-filtration. The bio-activity of partially purified protein was tested by MTT method and compared with standard IGF-I.

RESULTS: The amount of des (1-3) IGF-I expressed by pExSec1/2 (IGF-I) reached up to 19%-22% of the total soluble bacterial protein, which is about 7% higher than that of des (1-3) IGF-I expressed by pExSec1/IGF-I. The purity of recombinant des (1-3) IGF-I reached 49% and 82% respectively after the treatments by ultra filtration and gel-filtration. The result of MTT assay showed that the bio-activity of des (1-3) IGF-I after gel-filtration was about 77% of that of standard IGF-I at the same concentration.

CONCLUSION: The yield of recombinant des (1-3) IGF-I was increased about 7% by construction of expression plasmid with two copies of des (1-3) IGF-I gene, compared with only one copy of gene, preliminarily purified des (1-3) IGF-I showed relatively high biological activity, which was about 77% of that of standard IGF-I.

Keywords: Insulin-like growth factor I; Gene expression; Ultrafiltration; Chromatography, gel