摘要
为了探索Mayven蛋白的功能,进一步研究多发性硬化症的发病机制,我们通过PCR扩增得到Mayven4个不同长度的基因片段,包括片段P1(1-902 bp)、片段P2(1-523 bp)、片段P3(507-182 bp)和片段P4(887-1782bp),将这些片段克隆至大肠杆菌表达载体pGEX-4T-2,转化大肠杆菌BL21,并用IPTG诱导表达,对表达产物进行SDS-PAGE及western blot方法分析,采用GST蛋白纯化系统对融合蛋白进行纯化。实验结果显示我们成功构建了4个包含不同截短体的重组融合表达质粒GST-Mayven(P1、P2、P3、P4),诱导后表达得到4个截短体的Mayven融合蛋白,其表达形式均为可溶性表达,其相对分子质量与预期相符,纯化后得到各自的目的蛋白,为后续的Mayven蛋白的功能研究奠定了基础。
To understand the function of Mayven and investigate the pathogenesis of multiple sclerosis, the gene sequences of different truncated Mayven were amplified from the gene library of human brain. These truncated fragments, including fragment P1 （ 1-902 bp） ,fragment P2（ 1-523 bp）, fragment P3（507-182 bp） and fragment P4（887-1782 bp）, were cloned into pGEX-4T-2 vector to construct recombinant plasmids. The recombinant plasmids were transformed into E. coli BL21（DE3） and induced to express by IPTG. The expressed proteins were detected by SDS-PAGE and Western blot, and were purified by GST purifying system. The results showed that recombinant express vectors of different truncated GST-Mayven were successfully constructed and were expressed in soluble form protein induced by IPTG. The fusion proteins have good reactivity to GST antibody. The construction of recombinant express vectors of different truncated GST-Mayven lays a basis for further function study on Mayven.
出处
《生物医学工程学杂志》
EI
CAS
CSCD
北大核心
2008年第6期1401-1404,共4页
Journal of Biomedical Engineering
基金
重庆市科委重点项目资助(渝科发计字[2004]47号)
