摘要
为建立适用于小球藻（Chlorellasp.TLD6B）蛋白质组分析的双向电泳体系,该研究比较了TCA/丙酮沉淀法和Trisol提取法对小球藻蛋白的提取效果,不同pH梯度IPG胶条（pH3~10和pH4~7）、不同蛋白质上样量、不同聚焦程序对小球藻蛋白的分离效果。结果表明：（1）采用Trisol提取法可获得较高纯度蛋白,当选择24cm pH 3~10的线性IPG胶条时,上样量为500μg,聚焦80 000Vh效果最佳,可分辨蛋白点达726个;当选择24cm pH 4~7的线性IPG胶条时,上样量为1 000μg,聚焦80 000Vh效果最佳,可分辨蛋白点达1 230个。（2）该实验随机挑选了10个胶内蛋白点进行MALDI-TOF/TOF-MS鉴定分析表明,其中8个蛋白点鉴定成功,进一步说明Trisol提取法可适用于小球藻双向电泳分析。
In order to establish a two-dimensional gel electrophoresis（2D-E）protocol for proteomic study of desert chlorella（Chlorella sp.TLD6B）,we compared the performances of protein extraction methods from desert chlorella betweenTCA/acetone precipitation method and Trisol extraction.The chose pH gradient gel strips and protein loading volume were also optimized.Our results showed that：（1）trisol extraction could obtain high pure protein.The improved 2-DE system for desert chlorella was come out as follows：we distinguished 726 proteins with linear IPG strips of 24 cm in length with the pH ranges from 3to10,500μg loading volume,80000 Vh of IEF time.We distinguished 1230 proteins with linear IPG strips of 24 cm in length with the pH ranges from 4to 7,1 000μg loading volume,80 000 Vh of IEF time.（2）Ten randomly selected protein spots were identified by using MALDI-TOF/TOF-MS to confirm the establishment of two-dimensional electrophoresis protocol could be used for primary analysis of desert chlorella proteome.
出处
《西北植物学报》
CAS
CSCD
北大核心
2016年第9期1905-1911,共7页
Acta Botanica Boreali-Occidentalia Sinica
基金
国家自然科学基金(31460276)
关键词
小球藻
双向电泳
Trisol提取法
Chlorella
two-dimensional gel electrophoresis（2D-E）
Trisol extraction
作者简介
牟云（1992-），女，在读硕士研究生，主要从事可再生能源的开发与利用相关研究。E-mail：1171534413@qq．com
通信作者：高剑峰，教授，博士生导师，主要从事可再生能源的开发与利用相关研究。E-mail：jianfengg@shzu．edu．cn．
