Two-Dimensional Gel Electrophoresis and Pro-Q Diamond Phosphoprotein Stain-Based Plant Phosphoproteomics.
Two-Dimensional Gel Electrophoresis and Pro-Q Diamond Phosphoprotein Stain-Based Plant Phosphoproteomics.
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DOI：
10.1007/978-1-0716-1625-3_11
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发表时间：
2021
影响因子：
--
通讯作者：
Yuan Li;D. Ren
中科院分区：
文献类型：
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作者：
Yuan Li;D. Ren
Pro-Q钻石磷蛋白凝胶染色剂是一种荧光染色剂，用于检测聚丙烯酰胺凝胶中的磷酸化蛋白质，具有高灵敏度。在这里，我们描述了一个完整的程序，磷酸化蛋白质组学分析的组合Pro-Q钻石染色和双向凝胶电泳（2-DE）的拟南芥幼苗。工作流程包括总蛋白制备、2-DE蛋白分离、第二维凝胶染色、磷蛋白检测和用于基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）分析的肽制备。使用该方法可以检测大约300种磷蛋白。
Pro-Q diamond phosphoprotein gel stain is a fluorescent stain to detect phosphorylated proteins in polyacrylamide gels with high sensitivity. Here, we describe an entire procedure for phosphoproteomics analysis ofArabidopsisseedlings by a combination of Pro-Q diamond stain and two-dimensional gel electrophoresis (2-DE). The workflow involves total protein preparation, protein separation by 2-DE, the second-dimensional gel staining, phosphoproteins detection, and peptides preparation for matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis. Approximately 300 phosphoproteins can be detected using this method.