人白血病抑制因子基因在毕赤酵母中的表达

[Expression of human leukemia inhibitory factor gene in Pichia pastoris].

作者信息

Wang Q, Yang L, Quyang J, Long Q, Wang X

机构信息

Biopharmaceutical Center, State Key Laboratory for Biological Control, Zhongshan University, Guangzhou 510275, China.

出版信息

Wei Sheng Wu Xue Bao. 2001 Oct;41(5):582-6.

PMID:12552806

Abstract

The 575 bp artificial synthesized human leukemia inhibitory factor (hLIF) gene was cloned into vector pPICZ alpha A and the constructed expression vector pPICZ alpha A-hLIF was linearized by SacI and transformed into Pichia pastoris X-33. After the screening for Mut phenotype and the PCR analysis of the transformants, hLIF gene expressed in Pichia pastoris. SDS-PAGE and Western blot analysis of the culture supernatants showed that hLIF gene could express as 58.5 kD protein with the desired immunogenicity. Density scanning of the SDS-PAGE gel revealed that the targeted protein accounted for 32.8% of the total protein in the supernatants. The expressed products can inhibit the clone formation of murine's teratoma cells.

摘要

将575 bp人工合成的人白血病抑制因子（hLIF）基因克隆到载体pPICZαA中，构建的表达载体pPICZαA-hLIF经SacI线性化后转化到毕赤酵母X-33中。经过Mut表型筛选和转化子的PCR分析，hLIF基因在毕赤酵母中表达。对培养上清液进行SDS-PAGE和Western blot分析表明，hLIF基因可表达为具有所需免疫原性的58.5 kD蛋白。SDS-PAGE凝胶的密度扫描显示，目标蛋白占上清液中总蛋白的32.8%。表达产物可抑制小鼠畸胎瘤细胞的克隆形成。