山葡萄花序蛋白质双向电泳技术体系的建立

【目的】为开展山葡萄花序蛋白质组学研究。【方法】以山葡萄(Vitis amurensis Rupr)花序为试材,通过对蛋白质提取方法、裂解液成分、胶条pH梯度的改进,建立了适于山葡萄花序蛋白质组学研究的最佳双向电泳技术体系。【结果】采用酚提取法提取花序蛋白质,用含有硫脲和DTT的裂解液Ⅱ裂解,选用17 cm pH 4～7的IPG胶条在适宜的双向电泳条件下分离,可以获得分辨率高、背景清晰且重复性好的双向电泳图谱,可以检测出1 088个清晰的蛋白点。【结论】酚提取法适于山葡萄花序蛋白质的提取,硫脲和DTT可增强花序蛋白质的溶解性。

Establishment of two-dimensional electrophoresis platform for Vitis amurensis flower

【Objective】This work was performed to start the study of proteomic research using Vitis amurensis flower tissue.【Method】Vitis amurensis flower tissue proteome was extracted by two protein extraction protocols and lysis buffers,and various types of gel strips were evaluated.【Result】The results showed that phenol protein extraction method is superior than the TCA-acetone.The lysis bufferⅡwith thiourea and DTT is efficient in resolubilizing the protein pellet.17cm immobilized pH gradient strips with pH 4 to 7 could perform well in terms of protein separating in the first dimension.Finally,1 088 total protein spots were detected in the 2-DE gel map,which suggested that an efficient 2-DE platform with high repeatability and resolution for Vitis amurensis flower had been established.【Conclusion】Phenol protein extraction method was suitable for Vitis amurensis flower protein extract,thiourea and DTT enhanced flower protein solubility.