【摘要】 用重组DNA技术构建了病毒ki-ras表达质粒,用Eco R Ⅰ和Bam H Ⅰ酶解pUC-9DNA,小牛肠碱性磷酸酶脱磷酸,病毒ki-ras编码基因来源于Hi-Hi-3质粒DNA的Sat Ⅱ和Eco R Ⅰ酶切的0.6kb片段,用连接酶和DNA聚合酶Klenow片段将两个片段连接,转化大肠杆菌JM103,免疫筛选表达质粒,从156个转化克隆中得到两株表达质粒,其中一株pKras83的p21蛋白表达量为细菌总蛋白量的10％。更多还原

【Abstract】 In order to further investigate the biological and transforming activities of p21 and the possibility of using antibody against p21 for early diagnosis of cancer, we constructed a v-ki-ras expression plasmid with pAUC9 vector. The total cellular protein was analyzed by SDS-PAGE followed by staining with Coomassie blue. We found that there was a unique band, compared with the control, at Dalton 24 000. The same protein band gave a strong signal after Western transfer and reaction with 125 I labeled monoclonal anti-p21 antibody. The protein should be a fused protein of expressed p21 and a small pep-tide from lacZ, and composed more than 10% in total bacterial proteins. Preparation of antiserum is now in progress.更多还原