重组人B淋巴细胞刺激因子基因原核表达载体的构建及鉴定

Construction and Assessment of Gene Prokaryotic Expression Vector of Recombinant Human B Lymphocyte Cell Activating Factor

目的构建人BLyS基因原核表达载体。方法采用RT-PCR方法,从人外周血淋巴细胞的总cDNA中扩增得到474bp的人BLySDNA片段,再用BamHI和EcoRI双酶切后,克隆到原核表达载体pGEX-5X-3中,用限制性内切酶酶切分析和DNA序列分析鉴定重组质粒。结果BLyScDNA已经正确克隆到原核表达载体pGEX-5X-3中。结论本实验为研制抗人BLyS单克隆抗体来研究人BLyS与自身免疫性疾病的关系奠定了基础。

Objective To construct human BLys gene prokaryotic expression vector. Methods To amplified total cDNA of human peripheral lymphocyte cell and to get 474 bp fragment human BLyS DNA. Then it was cut by BamHI and EcoRI enzyme and was cloned into pGEX- 5X- 3 which is prokaryotic expression vector. The recombinant vector was analyzed and assessed by the means of restriction analysis and DNA sequence analysis. Results BLyS cDNA has been cloned into pGEX- 5X- 3 correctly. Conclusion This experiment established the foundation to study the relationship between human BLys and antoimmune disease by the means to develop anti human BLyS monoclonal antibody.