中肋骨条藻蛋白质双向电泳样品制备方法的比较

比较了裂解液法和直接三氯乙酸(TCA)-丙酮沉淀法对中肋骨条藻Skeletonema costatum 蛋白双向电泳的提取效果并优化了提取条件,结果表明:裂解液-TCA-丙酮沉淀法在去除胞内干扰物质方面,较裂解液-超滤管法和裂解液-试剂盒法的效果都好。 裂解液-TCA-丙酮沉淀法和直接TCA-丙酮沉淀法都能取得背景干净、蛋白点清晰的双向电泳图谱,但后者的双向电泳图谱聚焦更完全,在进一步优化条件后(即蛋白沉淀12 h并增加超声波洗涤过程),可作为提取中肋骨条藻蛋白的最适方法。

Comparison of protein sample preparation methods of two-dimensional electrophoresis for Skeletonema costatum

Lysis buffer method and trichloroacetic acid ( TCA)-acetone precipitation method both are the common methods in algal protein abstraction. We compared the 2-DE maps of these two protein extraction methods to find out the best protocol for Skeletonema costatum. As a result,the Lysis buffer-TCA-acetone method had better effect than the other two lysis buffer methods (lysis buffer-microcon &. lysis buffer-2-D clean-up kit) in removing the intracellular interferential factors, such as salt, nucleic acid, polyphenol and polysaccharide. The lysis buffer-TCA-acetone method and straight TCA-acetone precipitation method both represented clean background and clear protein dots in 2-DE maps while the later method showed better isoelect rofocusing results. We optimized the straight TCA-acetone precipitation methods. The TCA-acetone precipitation with 12 hours precipitation and a following ultrasonic cleaning process method was confirmed to be the appropiate sample preparation method for S. costatum for two-dimensional electrophoresis.