| 摘要: |
| 以杂色鲍(Haliotis diversicolor)血淋巴为材料进行了双向电泳研究。首先, 确定了以丙酮沉淀法和RC DC 蛋白定量试剂盒为基础的蛋白制备与定量方法; 其次, 对样品前处理方法进行了摸索, 通过500 r/min 的低速离心方法分离了血细胞和血浆组分, 通过35 000 r/min 超速离心2 h 除掉了部分高丰度的血蓝蛋白, 降低了高分子量区域的血蓝蛋白背景; 进一步使用18 cm pH 3-10 非线性胶条和标准化的电泳程序, 对不同上样量的电泳银染结果进行了比较, 获得了良好的双向电泳图谱; 最终选取肌动蛋白、原肌球蛋白、胶原蛋白和血蓝蛋白4 个蛋白点成功进行了质谱鉴定。 |
| 关键词: 杂色鲍(Haliotis diversicolor) 血淋巴 血浆 双向电泳 蛋白质组 |
| DOI： |
| 分类号: |
| 基金项目:现代农业产业技术体系建设专项资金-贝类产业技术体系科学家岗位专项经费项目(CARS-48); 中央级公益性科研院所基本科研业务费专项资金项目(2010TS16) |
|
| Two-dimensional electrophoresis and mass spectrometry analysis of Haliotis diversicolor hemolymph |
|
|
| Abstract: |
| Hemolymph of abalone was used as the material for the two-dimensional electrophoresis research. At first, protein preparation and quantification method based on the acetone precipitation and the RC DC Protein Assay Kit was successfully set up. Then pre-treatment methods with 500 r/min low-speed centrifugation and 35000 r/min ultracentrifugation were used for the separation of hemocyte and blood plasma or reducing the high molecular weight region’s background. Furthermore, non-linear strips (18 cm, pH 3-10) and standardized electrophoresis protocols were established to evaluate different sample volumes with silver stain. Good two-dimensional electrophoresis maps were finally acquired, and four protein spots in the map were successfully identified by mass spectrometry, which are actin, tropomyosin, collagen and hemocyanin. |
| Key words: abalone hemolymph blood plasma two-dimensional electrophoresis proteome |
