期刊文献+

红鳍东方鲀FoxO1基因的原核表达及纯化 被引量:3

Prokaryotic expression and purification of FoxO1 gene from torafugu Takifugu rubripes
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摘要 采集红鳍东方鲀脂肪组织提取其总RNA,采用RT–PCR方法扩增和克隆红鳍东方鲀FoxO1基因的ORF,并构建其原核表达载体p ET–32/FoxO1,在大肠杆菌Rosetta(DE3)进行表达,通过IPTG诱导表达,对重组FoxO1蛋白进行可溶性分析、纯化及鉴定,结果表明:重组质粒p ET32a/FoxO1被成功构建;用IPTG进行诱导表达,融合蛋白主要以可溶蛋白形式存在,能与抗His标签的兔多克隆抗体发生特异性反应;纯化出了融合表达蛋白,获得了相对分子质量约为110 000的重组蛋白。 The ORF of FoxO1 gene was amplified by reverse transcription-polymerase chain reaction(RT–PCR) from total RNA extracted from adipose tissues of torafugu Takifugu rubripes. PET–32a/Sirt1,the prokaryotic expression vector,was constructed and transformed into Escherichia coli Rosetta(DE3) for FoxO1 expression. Through IPTG inducting expression,the induced fusion protein was purified and verified by SDS–PAGE and Western Blot. As a result,the full-length open reading frame of torafugu FoxO1 c DNA was successfully cloned,and the recombinant vector was constructed. The expressed fusion proteins induced by IPTG were soluble protein. The fusion proteins with tag His were also purified,and the molecular weight of the recombinant protein was about 110 000.
出处 《湖南农业大学学报(自然科学版)》 CAS CSCD 北大核心 2016年第1期81-84,共4页 Journal of Hunan Agricultural University(Natural Sciences)
基金 河南省科学技术厅科技攻关计划项目(122102310196) 河南省教育厅自然科学研究计划项目(12A150019)
关键词 红鳍东方鲀 FoxO1基因 原核表达 蛋白纯化 Takifugu rubripes FoxO1 gene prokaryotic expression protein purification
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