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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。
Objective: To establish a two-dimensional gel electrophoresis pattern of JAR / MTX proteome in human choriocarcinoma cell line. Methods: JAR / MTX cells were cultured in RPMI-1640 medium containing 10% fetal bovine serum. After trypsinization, cells were harvested and cells were lysed to complete lysis. After centrifugation, the supernatant was subjected to two-dimensional gel electrophoresis and test MASL bright blue staining, the application of PDQuest image analysis system for gel analysis, from the gel map to select a good separation of protein spots, using quadrupole - time of flight mass spectrometry (Q-TOF-MS) and database search to identify proteins . Results: A two-dimensional gel electrophoresis pattern with clear background, good resolution and good reproducibility was obtained. (810 ± 45) protein spots with good reproducibility and resolution were obtained on 17cmpH3 ~ 10IPG strips, The matching rate of protein spots was 80.2%; a protein spot was identified by mass spectrometry and database search. Conclusion: The JAR / MTX two-dimensional gel electrophoresis reference map of human choriocarcinoma cell line was established, which laid the foundation for its further study of proteomics.