用3种方法提取中华绒螯蟹卵巢总蛋白效果的比较

为了探寻适于中华绒螯蟹Eriocheir sinensis卵巢蛋白质组学研究的总蛋白最佳提取方法,采用RC DC Protein Assay检测蛋白浓度、SDS-PAGE分析蛋白组成等,比较分析了用裂解液法Ⅰ、裂解液法Ⅱ和TCA/丙酮沉淀法提取中华绒螯蟹卵巢蛋白的效果。结果表明:应用3种方法提取的卵巢总蛋白浓度分别为15.59、13.05、0.65 mg/m L,用裂解液法提取的效果最佳,而TCA/丙酮沉淀法不适合用于中华绒螯蟹卵巢总蛋白的提取;SDS-PAGE电泳图谱进一步显示,用裂解液法Ⅱ提取的蛋白样品组成较为丰富,且重复性好,比较适合后续蛋白质组学研究。该研究结果可为中华绒螯蟹卵巢发育蛋白质组学研究提供参考。

Comparative extraction of total proteins in ovary of Chinese mitten handed crab Eriocheir sinensis by different methods

The extraction effects of two lysis buffer methods and trichloroacetic acid/acetone precipitation method on total proteins in ovary of China mitten crab handed Eriocheir sinensis were comparatively studied for the pro-teomics by RC DC protein assay in order to search for the optimal extraction method to detect the protein level, and the protein composition was analyzed by SDS-PAGE. The results showed that the protein concentration was 15. 59 mg/mL and 13. 05 mg/mL by lysis buffer methods and 0. 65 mg/mL, by trichloroacetic acid/acetone precipitation method, indicating better by lysis buffer methods than by trichloroacetic acid/acetone precipitation. The SDS-PAGE maps analysis revealed that the sample extracted by lysis buffer methodⅡ showed more protein bands, and had high reproducibility. The findings indicate that lysis buffer methodⅡis the better method for the proteomics of ovary proteins in China mitten handed crab, providing a reference for proteomics of China mitten handed crab.