| 质谱蛋白质组学研究中草莓雌蕊蛋白提取方法优化 |
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| 引用本文： | 裘劼人,柴伟国,童建新,周历萍,王淑珍.质谱蛋白质组学研究中草莓雌蕊蛋白提取方法优化[J].浙江农业学报,2020,32(12):2186. |
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| 作者姓名： | 裘劼人 柴伟国 童建新 周历萍 王淑珍 |
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| 作者单位： | 杭州市农业科学研究院,浙江 杭州 310024 |
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| 基金项目： | 杭州市农业科研自主申报项目(20191203B53);杭州市农科院科技创新基金(2020HNCT-21) |
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| 摘 要： | 为筛选适用于质谱蛋白质组学研究的草莓雌蕊蛋白提取方法,通过在传统酚提法中加入氯仿提取步骤建立了一种新的酚氯仿提取方法,并对三氯乙酸/丙酮沉淀法(方法Ⅰ)、酚提法(方法Ⅱ)和酚氯仿法(方法Ⅲ)提取草莓雌蕊蛋白的3种方法的适用性进行了评估。总蛋白提取得率、总蛋白鉴定通量、非共有蛋白亚细胞位置和亲疏水性分布4个方面的分析结果表明,酚氯仿法的蛋白提取效率最高,蛋白提取最全面。方法Ⅰ、方法Ⅱ、方法Ⅲ的蛋白提取得率分别为0.20%、0.12%和0.27%,总蛋白鉴定量分别为5 170、5 345和5 697。韦恩图分析显示,方法Ⅲ交集到了方法Ⅰ和方法Ⅱ的大部分非共有蛋白。亚细胞定位分析显示,方法Ⅰ和方法Ⅲ的非共有蛋白主要分布在细胞核中,而方法Ⅱ则主要分布在细胞质中,表明3种方法具有不同的亚细胞偏好性。亲疏水性分析显示,方法Ⅰ亲水性蛋白比例最高但疏水性蛋白提取不足,方法Ⅱ则亲水性蛋白提取较少,方法Ⅲ兼顾了亲疏水性蛋白的提取。试验结果表明,3种方法中方法Ⅲ为草莓雌蕊的最优提取方法。
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| 关 键 词： | 草莓 雌蕊 蛋白质组学 提取方法 |
| 收稿时间： | 2020-07-13 |
Optimization of strawberry gynoecium protein extraction method for mass spectrum-based proteomic study |
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| QIU Jieren,CHAI Weiguo,TONG Jianxin,ZHOU Liping,WANG Shuzhen.Optimization of strawberry gynoecium protein extraction method for mass spectrum-based proteomic study[J].Acta Agriculturae Zhejiangensis,2020,32(12):2186. |
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| Authors: | QIU Jieren CHAI Weiguo TONG Jianxin ZHOU Liping WANG Shuzhen |
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| Institution: | Hangzhou Academy of Agricultural Sciences, Hangzhou 310024, China |
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| Abstract: | In order to screen a strawberry gynoecium protein extraction method for mass spectrum (MS)-based proteomics studies, a new method named as phenol-chloroform extraction was established by adding a chloroform extraction step to the traditional phenol extraction method, and the application on extracting strawberry gynoecium proteins of the three methods: trichloroacetic acid/acetone precipitation (method Ⅰ), phenol extraction(method Ⅱ) and phenol-chloroform extraction (method Ⅲ) were evaluated. The analysis results of the total protein yield, sum of protein identification, subcellular location and hydrophilic-hydrophobic distribution of non-shared proteins showed that, the phenol-chloroform method was the most efficient and comprehensive for protein extraction. The protein yields of method Ⅰ, method Ⅱ and method Ⅲ were 0.20%, 0.12% and 0.27%, respectively; and the sum of protein identified were 5 170, 5 345, and 5 697, respectively. The Venn diagram analysis showed that method Ⅲ intersected most of the non-shared proteins of method Ⅰ and method Ⅱ. Subcellular location analysis showed that non-shared proteins of method Ⅰ and method Ⅲ were mainly distributed in the nucleus, whereas those of method Ⅱ were mainly distributed in the cytoplasm, indicating that the three methods had different subcellular preferences. Hydrophilic-hydrophobic analysis showed that method Ⅰ had the highest proportion of hydrophilic protein but insufficient hydrophobic protein extraction, while method Ⅱ had insufficient extraction of hydrophilic protein, and method Ⅲ took into account the extraction of both hydrophilic and hydrophobic proteins. According to the results, method Ⅲ should be the optimal protein extraction method for strawberry gynoecium. |
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| Keywords: | strawberry gynoecium proteomics extraction methods |
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