Comparison of protein extraction methods and two-dimensional electrophoresis analysis in sacred lotus seeds
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摘要: 莲（Nelumbo nucifera Gaertn.）不仅是重要的水生蔬菜作物之一，而且是进行基础研究的好材料。本文采用4种蛋白质提取方法（新型TCA/丙酮法、传统TCA/丙酮法、改良的Tris-HCl法、Tris-饱和酚法）并结合双向电泳技术，对莲子蛋白质提取方法进行筛选与优化。双向电泳实验结果显示，所得蛋白质图谱与莲种子蛋白质组成分布特点一致。通过PDQuest软件分析表明，新型TCA/丙酮法适用于莲子叶和胚芽组织的双向电泳蛋白质提取，而传统TCA/丙酮法则适用于莲胚轴组织双向电泳的蛋白质提取。研究结果为进一步利用质谱进行莲子蛋白质组研究奠定了基础。Abstract: Nelumbo nucifera Gaertn. (sacred lotus) is not only an important aquatic vegetable crop, but also good material for basic scientific research. The extraction of sacred lotus seed proteins was optimized using four protein extraction methods (new TCA/acetone, traditional TCA/acetone, improved Tris-HCl, and Tris-saturated phenol methods) combined with two-dimensional electrophoresis (2-DE) technology. The 2-DE analysis showed that the resulting protein profile/map exhibited consistency with the composition and distribution of the sacred lotus seed proteome in ‘Tielian’. Analysis by PDQuest software showed that the new TCA/acetone method was suitable for 2-DE protein extraction of lotus cotyledons and plantules, whereas the traditional TCA/acetone method was suitable for the 2-DE protein extraction of hypocotyls. This work lays a foundation for further research on the sacred lotus seed proteome by mass spectrometry.
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