Tian S, Liu L, Feng B
Norman Bethune University of Medical Sciences, Changchun 130021.
Zhonghua Xue Ye Xue Za Zhi. 1997 Dec;18(12):634-7.
To obtain human thrombopoietin (Tpo)mutant cDNA encoding mature peptide N terminal 1 approximately 196 amino acids and investigate its expression in E. coli JM109.
Polymerase chain reaction and DNA recombination techniques were employed. PCR product was inserted into pUC19 vector and sequenced and then cloned into expression vector pMAL-c2.
E. coli JM109 cells with plasmid pMAL -MBP/TpoM were induced by IPTG for 4-5 hours. SDS-PAGE analysis showed that MBP/TpoM molecular weight is about 63KD. Scanning analysis indicated that expressed protein accounts up to 37% of total E. coli proteins.
The Tpo mutant of interest is successfully expressed in E. coli JM109 cells.
获取编码成熟肽N端约196个氨基酸的人血小板生成素（Tpo）突变体cDNA，并研究其在大肠杆菌JM109中的表达。
采用聚合酶链反应和DNA重组技术。将PCR产物插入pUC19载体并测序，然后克隆到表达载体pMAL-c2中。
用异丙基-β-D-硫代半乳糖苷（IPTG）诱导携带质粒pMAL-MBP/TpoM的大肠杆菌JM109细胞4至5小时。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）分析表明，MBP/TpoM的分子量约为63千道尔顿（KD）。扫描分析表明，表达的蛋白占大肠杆菌总蛋白的37%。
目的Tpo突变体在大肠杆菌JM109细胞中成功表达。
