【摘要】 目的:建立人绒癌耐药细胞株JAR/MTX蛋白质组双向凝胶电泳图谱。方法:用含10%胎牛血清的RPMI-1640培养液培养JAR/MTX细胞,胰酶消化后收集细胞,加入细胞裂解液使细胞充分裂解,离心后取上清液进行双向凝胶电泳和考马斯亮蓝染色,应用PDQuest图像分析系统进行凝胶图谱分析,从凝胶图中选取1个分离较好的蛋白质点,应用四极杆-飞行时间质谱(Q-TOF-MS)和数据库搜索鉴定蛋白质。结果:获得了背景清晰、分辨率和重复性较好的双向凝胶电泳图谱;在17cmpH3～10IPG胶条上可分离到(810±45)个重复性及分辨率均较好的蛋白位点,蛋白斑点的匹配率为80.2%;1个蛋白点通过质谱分析和数据库检索得到了初步的鉴定。结论:建立了人绒癌耐药细胞株JAR/MTX双向凝胶电泳参考图谱,为其蛋白质组学的进一步研究奠定了基础。更多还原

【Abstract】 Objective:To establish two-dimensional gel electrophoresis profiles of proteome from drug-resistant choriocarcinoma cell line JAR/MTX.Methods:JAR/MTX cells were cultured in RPMI1640 culture medium containing 10% fetal bovine serum,then harvested after digestion by trypsin,and lysed in lysis buffer.After centrifugation,supernatant was separated by isoelectric focusing (IEF),equilibrated and run on SDS polyacrylamide gel electrophoresis (SDS-PAGE),then stained by Coomassie brilliant blue.A well-seperated protein spot was selected from the gel by PDQuest analysis system.Hybrid Quadrupole Time of Flight Mass Spctrometer (Q-TOF-MS),peptide mass fingerprinting (PMF) and database searching were used to identify the protein spot.Results:Clear,well-resolved,reproducible two-dimensional gel electrophoresis profiles of proteome in JAR/MTX cells were obtained.(810±45) protein spots were detected in the two-dimensional gel electrophoresis profiles of proteome on 17 cm IPG strip (pH 3～10) of JAR/MTX cell proteome separated.A match rate of 80.2% of the spots between gels was obtained.One of these proteins was identified by mass spectrometry and database queries.Conclusion:The two-dimensional gel electrophoresis profiles of proteome from JAR/MTX cells are established and lay the foundation of further proteomics research of JAR/MTX cells.更多还原