RPTPα-D2的克隆表达及其多克隆抗体的制备

发布时间：2018-06-26 21:38

本文选题：RPTPα + 抗体　；参考：《吉林大学》2006年硕士论文

【摘要】：受体型蛋白酪氨酸磷酸酶α(Reacptor Protein Tyrosine Phosphataseα,RPTPα)是一种广泛表达的跨膜受体型蛋白酪氨酸磷酸酶(ProteinTyrosine Phosphatases ,PTPs),能够在体外对c-Src进行去磷酸化,从而增强c-Src的激酶活性。因此,RPTPα特异性抑制剂能抑制c-Src家族激酶活性,在治疗由c-Src家族激酶活性增高引起的肿瘤疾病中具有重要的应用价值。我们以pT7-RPTPα的序列为模板,克隆了RPTPα的细胞内结构域RPTPα-D2,构建了原核表达载体pGex-His-C3-RPTPα-D2质粒,转化大肠杆菌BL21 Codon plus,进行融合蛋白质GST-RPTPα-D2的可溶性高效表达,并利用Sepharose 4B亲和层析柱纯化该融合蛋白。使用凝血酶消化除去GST蛋白,并用纯化后的RPTPα-D2蛋白免疫家兔,制备了RPTPα-D2的多克隆抗体,并对其进行纯化,这为进行RPTPα的细胞定位和筛选RPTPα特异性抑制剂奠定了基础。
[Abstract]:Receptor protein tyrosine phosphatase 伪 (RPTP 伪) is a widely expressed protein tyrosine phosphatases (PTPs), which can dephosphorize c-Src in vitro and enhance the activity of c-Src kinase. Therefore, RPTP 伪 -specific inhibitors can inhibit the activity of c-Src family kinases, and have important application value in the treatment of tumor diseases caused by increased activity of c-Src family kinases. Using the sequence of pT7-RPTP 伪 as template, we cloned the intracellular domain RPTP 伪 -D2 of RPTP 伪, constructed the prokaryotic expression vector pGex-His-C3-RPTP 伪 -D2, transformed it into E. coli BL21 Codon plus. and expressed the fusion protein GST-RPTP 伪 -D2. The fusion protein was purified by Sepharose 4 B affinity chromatography. Using thrombin digestion to remove GST protein and immunizing rabbits with purified RPTP 伪 -D2 protein, the polyclonal antibody of RPTP 伪 -D2 was prepared and purified, which laid a foundation for the cellular localization of RPTP 伪 and the screening of RPTP 伪 specific inhibitors.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R392

【相似文献】