摘要
为了获取高质量的西藏巨柏叶片蛋白质,分别采用酚抽提法、TCA/丙酮法和改良的TCA丙酮法3种方法对西藏巨柏叶片进行总蛋白的提取,同时利用蛋白试剂盒(The 2-D Quant Kit)对蛋白质的浓度和纯度进行了分析测定,并通过蛋白质斑点数量及分布特征,最后确定了最佳蛋白提取方法。结果表明:对样品研磨的精细程度、丙酮清洗沉淀次数等操作细节均能影响提取结果,同时采用改良的TCA丙酮法提纯后的结果最佳。说明改良的TCA丙酮法可应用于西藏巨柏叶片后续的蛋白质组学研究。
In this study, to extract high-quality leaves protein of Cupressus gigantea, total leaves protein was extracted using three methods including Phenol extraction method, TCA-acetone method and modified TCA-acetone method. The concentrates and purity of the proteins were determined by The 2-D Quant Kit, and then two-dimensional gel electrophoresis was conducted in the test. The optimal method of protein extraction was determined through comparing the protein concentration, the number and distribution characteristics of protein spots. The results showed that some operational details could affect the extraction results, including the grinding degree of samples, the number of precipitation washed with acetone, and so on. The electrophoresis result was optimal through using the modified TCA-acetone method for the purification. The result indicates that the modified TCA-acetone method can be applied to the further study of leaves proteomics of Cupressus gigantea.
引文
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