黄瓜悬浮细胞蛋白质组双向电泳分析技术

为建立适于黄瓜悬浮细胞蛋白质组分析的双向电泳体系,对黄瓜悬浮细胞蛋白质双向电泳分析所采用的胶条pH范围、样品制备方法、裂解液配方及分离胶浓度等参数进行研究。结果表明,采用pH范围为4～7的IPG胶条,直接裂解后丙酮沉淀法制备黄瓜悬浮细胞蛋白质,裂解液为8mol/L尿素、2mol/L硫脲、2%IPG Buffer、4%CHAPS、1%TBP、65mmol/L DTT、2mmol/L EDTA、0.001%溴酚蓝和1%鸡尾酒,分离胶浓度为11%,可获得蛋白质点分离清晰的双向电泳图谱。

Optimization of Two-dimensional Electrophoresis for Proteome of Cucumber Suspension Cultured Cells

To obtain suitable protocol for proteome of cucumber suspension cultured cells,we optimized the pH gradient of IPG strip,the method of protein extraction,reagent and concentration of lysis buffer and SDS-PAGE condition.Results showed pH 4~7 IPG strips,the extraction method acetone precipitation after lysis,lysis buffer containing 8 mol/L urea,2 mol/L thiourea,2% IPG Buffer,4% CHAPS,1% TBP,65 mmol/L DTT,2 mmol/L EDTA,0.001% bromophenol blue and 1% cocktail and 11% gel concentration were appropriate for the sample preparation of cucumber suspension cultured cells.