独行菜属植物种子总蛋白4种提取方法的比较  

Comparative Study of Four Methods of Total Protein Extraction from Seeds of Two Species of Lepidium Plants

作　　者：丁金鹏[1] 张娜[1] 李群[1] 

机构地区：[1]新疆大学生命科学与技术学院,乌鲁木齐830046

出　　处：《新疆农业科学》2017年第7期1305-1312,共8页Xinjiang Agricultural Sciences

基　　金：国家自然科学基因项目"新疆濒危植物高山离子芥抗冷相关功能基因组分析与关键基因克隆"(31160051);新疆维吾尔自治区重大科技专项"新疆早春短命植物与模式植物拟南芥的比较基因组学与功能基因组学研究"(200840102-40)~~

摘　　要：【目的】研究适用于抱茎独行菜和独行菜种子SDS-PAGE电泳和双向电泳的蛋白样品提取方法,为研究两种独行菜种子蛋白组学奠定基础。【方法】采用4种蛋白质提取方法提取种子总蛋白,测定和分析蛋白产量和SDS-PAGE电泳图谱。【结果】TCA/丙酮提取法获得蛋白量较多,杂质少,电泳条带多且清晰;裂解液提取法蛋白的产量最高,有拖尾且杂质多;Tris-HCl提取法和苯酚提取法的蛋白产量低,杂质少;裂解液提取法和苯酚提取法获得的蛋白在SDS-PAGE电泳时易产生高丰度蛋白的干扰。对比TCA/丙酮提取法和苯酚提取法获得的两种独行菜种子总蛋白SDS-PAGE图谱,发现有6条蛋白差异条带。【结论】TCA/丙酮提取法提取两种独行菜种子总蛋白效率高,纯度高且杂质少,可用于SDS-PAGE电泳和双向电泳分析;两种独行菜种子蛋白SDS-PAGE电泳的差异谱带可为甄别两种独行菜种子提供参考依据。【 Objective】 To explore the extraction methods of protein samples for SDS - PAGE and two - dimensional electrophoresis （2 - DE） of the stem, vegetable, and vegetable seeds, and lay the foundation for the study of seed protein proteomics of the two species of Lepidium plants.【 Method】 The efficiency of four protein extraction methods were compared and analyzed through protein purity detection and SDS - PAGE. [Result] Of all the four methods, TCA/acetone extraction method generated higher yield as well as highest resolution and clear estelectrophoresis bands in SDS - PAGE, lysis extraction method got the highest protein yield whereas with highest impurities, Tris - HC1 method and phenol method could obtain the highest purity but lowest protein yield, meanwhile protein extracted by lysis method and phenol method had the highest level of high - abundance proteins which could cause serious interference in further SDS - PAGE and 2 - DE analy-sis. We got 6 differential expressed bands after compared between two species of Lepidium plants by SDS - PAGE using seed total protein extracted by TCA/acetone extraction method and phenol method.【 Conclusion】 TCA/acetone seed total protein extraction method is an optimal sample preparation protocol for SDS - PAGE and 2 - DE. Besides, the differential bands of SDS - PAGE electrophoresis of two kinds of Lepidium seed pro-tein could provide reference for screening two seeds.

关 键 词：独行菜 抱茎独行菜 种子总蛋白 蛋白质提取 SDS-PAGE 

分 类 号：S188[农业科学—农业基础科学]