NaHCO3胁迫下刚毛柽柳叶片蛋白质差异表达谱的建立
Establishment of protein differential expression map of Tamarix hispida leaves under NaHCO3 stress
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摘要:目的为了研究刚毛柽柳响应盐胁迫的作用机制, 本研究以刚毛柽柳叶片为材料, 对叶片总蛋白质的提取方法进行优化, 探究出适用于双向电泳的刚毛柽柳叶片总蛋白质的提取方法, 并建立NaHCO3胁迫下刚毛柽柳叶片差异蛋白质的表达谱。方法本研究比较了改良Tris-三氯乙酸, 改良Tris-三氯乙酸+PVP40, 改良Tris-三氯乙酸+丙酮和植物蛋白提取试剂盒法4种提取植物蛋白质的方法提取刚毛柽柳叶片总蛋白质的差异, 并利用蛋白质双向电泳用技术研究NaHCO3胁迫后刚毛柽柳叶片蛋白质表达变化, 建立了盐胁迫下刚毛柽柳叶片差异蛋白表达谱。结果采用改良Tris-三氯乙酸法提取刚毛柽柳叶片蛋白质, 利用该方法可获得高质量的蛋白质样品, 所得到的双向电泳凝胶图谱中蛋白质分离效果好、图谱分辨率高, 通过ImageMaster 2D Platinum 7.0凝胶分析软件对NaHCO3处理0和12 h的双向电泳凝胶图谱进行分析, 获得了25个响应盐胁迫的差异表达蛋白质, 并分析了差异表达蛋白量的变化。结论研究结果表明, 利用改良Tris-三氯乙酸法提取刚毛柽柳叶片蛋白质适用于双向电泳体系, 该方法可以用于对刚毛柽柳叶片蛋白质组学的研究。本研究建立了NaHCO3胁迫下刚毛柽柳叶片差异蛋白质的表达谱, 为进一步分析刚毛柽柳耐盐分子机制奠定了基础。Abstract:ObjectiveIn order to study the mechanism of Tamarix hispida responsing to the salt stress, this study is committed to optimize the extraction method of total protein from the leaves of Tamarix hispida. The main purpose of this research is to explore the method of extracting total protein which is suitable for two-dimensional electrophoresis of Tamarix hispida leaves, and establish the differential protein expression map of Tamarix hispida leaves under NaHCO3 stress.MethodImproved Tris/TCA, improved Tris/TCA/PVP40, improved Tris/TCA/Acetone and plant protein extraction kit methods were used to study the optional extracting protein methods of Tamarix hispida leaves. Differential expression of proteins in Tamarix hispida leaves under NaHCO3 stress were studied using two-dimensional electrophoresis. And the protein differential expression map of Tamarix hispida leaves under salt stress was established.ResultThis research revealed that we can obtain the high purity of protein sample and the map of two-dimensional gel electrophoresis which has good separation and high resolution using Tris/TCA method. The two-dimensional gel electrophoresis maps of proteins which were treated with NaHCO3 for 0 h and 12 h were analyzed by ImageMaster 2D Platinum 7.0 gel analysis software. Twenty-five differentially expressed proteins were detected, which were responded to salt stress and the changes of differentially expressed proteins were analyzed.ConclusionThe results showed that the extraction of Tamarix hispida leaf protein by improved Tris/TCA method is suitable for two-dimensional electrophoresis system, and this method could be used to study the proteome of Tamarix hispida leaves. In this study, the differential protein expression map of Tamarix hispida leaves under NaHCO3 stress was established, and this result would be the foundation of the molecular mechanism of Tamarix hispida with salt tolerance for further analysis.
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