蛋白质双向电泳、印迹转移及蛋白质合成无细胞体系技术在小鼠腹水细胞EF-3研究中的应用

The Application of the Methods of Two Dimentional Gel Electrophoresis, Western Blotting and Cell-free System of Protein Synthesis in the Research in EF-3 Factor of H22a Cell

本文应用近年发展起来的生化技术－蛋白质双向电泳（其第一向为等电聚焦，第二向为ＳＤＳ凝胶电泳），将小鼠腹水细胞核粮体蛋进行了指纹分离，并利用蛋白质印迹转移（Ｗｅｓｔｅｒｎ ｂｌｏｔｔｉｎｇ），将转移后的硝酸纤维素膜与交联了碱性磷酸的第二抗体和抗酵缮母ＥＦ－３抗体反应，证实该核粮体蛋白含有ＥＦ－３同源片段，进而制备了蛋白质合成无细胞体系。通过测定ＰｏｌｙＵ指导下＾３Ｈ－ｐｈｅ掺入活力的免疫失活实验。

The ribosomal proteins of H22a cell,were separated with the method of two-D gel clectrophoresis(the first dimention is isoclcctric focus and the second is SDS PAGE).Then the Western blotting was used,the transferred nitrocellulose sheet was treated with antiycast EF-3 antibody and the second antibody bonded with alklincphosphoestcrasc.The result shows that the ribosomal proteins have a homologous fragment to yeast EF-3 factor.The cell-free system of protein synthesis was also established.By dcterming the activity of polyU directed 3H-phc intervention in immunodeactivitive experiment,it is primarily confirmed that this fragment is the essential for the protein biosynthesis in H22a cell.