奇异变形杆菌蛋白质双向电泳条件的优化及评价

目的：优化针对奇异变形杆菌蛋白质组学研究的双向电泳(2-DE)条件及其相关技术体系,阐明样品处理过程中超声功率、振摇时间和上样量对2-DE图谱的影响。方法：选取1株奇异变形杆菌，分别采用超声功率为130 W和80 W超声法裂解菌体提取总蛋白，将2种方法提取的蛋白在同一条件下进行等电聚焦和聚丙烯酰胺凝胶电泳(SDS-PAGE);超声功率为80 W提取的奇异变形杆菌总蛋白，分别取上样量1和2 mg，在同一条件下17 cm胶条上进行双向电泳。结果：超声功率为130 W处理的样本得到的2-DE图谱中蛋白斑点较少，而超声功率为80W处理的样本蛋白点多、清晰且重复性好。上样量为2 mg得到的图谱中横纵条纹明显，蛋白斑点分离度较差，而上样量为1 mg时横纵条纹较少，蛋白斑点分离度较好。结论：采用低功率超声且较少上样量处理样品能得到较好的2-DE图谱。
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Optimization and evaluation of two-dimensional electrophoresis conditions of Proteus mirabilis

Objective To optimize the conditions of two-dimensional electrophoresis(2-DE) and its related techniques in research on proteomics of Proteus mirabilis,and to elucidate the effects of the supersonic power,the agitate time and the loading amount of proteins on 2-DE maps.Methods One strain of Proteus mirabilis was chosen,and its proteins were extracted by sonicating on a sonicator fitted with a microprobe.and the supersonic powers were 130 W and 80 W.The same condition of immobilized pH gradient isoelectric focusing and SDS-PAGE was used to separate the proteins obtained from the two ways.1 and 2 mg of samples of the total proteins obtained from Proteus mirabilis by 80 W supersonic power were collected for 2-DE on 17 cm IPG strips under the same condition.Results The 2-DE map of the proteins obtained by 130 W supersonic power was clear,repeatable and had more protein spots than that obtained by 80 W supersonic power.The 2-DE map with sample amount of 1 mg exhibited less vertical and horizontal stripes than that of 2 mg,which appeared good repeatability.Conclusion Adopting lower supersonic power and a few of samples can get better 2-DE map.