TIR突变的hbFGF在原核系统中高效的表达  

The study of TIR-modified hbFGF being highly expressed in prokaryotic system

作　　者：陈小佳[1] 林剑[1] 汪矩[1] 孙奋勇[1] 吴健虹[1] 洪岸[1] 

机构地区：[1]暨南大学生物工程研究所,广东广州510632

出　　处：《暨南大学学报（自然科学与医学版）》2003年第5期110-116,共7页Journal of Jinan University(Natural Science & Medicine Edition)

基　　金：国家九五科技攻关重点项目(96-C02-01-03);国家863高科技项目(Z18-03-29);广州市科委重点科技攻关项目(99-Z-003-01)

摘　　要：　目的:为获得非融合高表达hbFGF的大肠杆菌表达系统;方法:设计PCR引物,将hbFGF的5'端前10个密码子突变,优选大肠杆菌较偏好的密码子并降低G+C含量,通过双酶切法将突变后的hbFGF克隆入pBV220质粒载体中,筛选出重组子,再通过温控法对重组菌体进行诱导,分析表达情况,并进一步纯化和测活;结果:bFGF在该系统中高水平表达,表达量超过30%,其中绝大部分形成包涵体,经过包涵体变复性等纯化步骤后测活性,证明有生物活性;结论:TIR区域的基因突变对hbFGF在pBV220中高效表达起到关键作用.Aim:To obtain highly-expressed prokaryote system of non-fusion hbFGF. Methods: As a template, hbFGF gene was modified for changing 10 codons of 5′ end with Ecoli preference codons by PCR. Using pBV220 as vector, the cloned hbFGF was expressed in DH5α. The target protein was extracted and then followed by cation-exchange chromatography and gel filtration. And its bioactivity detected by MTT assay. Results: hbFGF was highly expressed up to 30% in this prokaryote system. Most was inclusion body. The target protein have the same bioactivity with standard bFGF. Conclusion: Modification of hbFGF TIR is an effective means to increase nonfusion expression rate of hbFGF in prokaryote system.

关 键 词：翻译起始区 人碱性成纤维细胞生长因子 大肠杆菌 非融合 高效表达 

分 类 号：Q789[生物学—分子生物学]