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作者	单位
范锋贵，张晓科，任万杰，王晓龙，叶石，付晓洁，王宏礼，朱建楚	（西北农林科技大学农学院/国家小麦改良中心杨凌分中心，陕西杨凌 712100 ）

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中文摘要:

水分胁迫应答蛋白的表达与小麦品种的抗旱性密切相关。为了明确与抗旱相关的水分胁迫D-应答蛋白的表达特点及蛋白组成，对小麦品种晋麦47幼苗进行不同供水量处理，应用SDS-PAGE、纳升级液相色谱-电喷雾串联质谱联用技术（Nano LC-MS/MS），分析了水分胁迫D-应答蛋白条带表达量的变化和蛋白组成。SDS-PAGE检测结果表明，水分胁迫D-应答蛋白条带在-0.5 MPa PEG-6000水溶液胁迫下，6 h出现可见表达，胁迫至48 h时表达量最大，之后，此蛋白条带的表达量又逐渐下降；幼苗胁迫48 h后恢复正常供水，复水72 h后该蛋白消失。切取正常供水和胁迫处理两个条件下SDS-PAGE胶差异表达蛋白条带，经质谱分析分别获得两个阳性结果，且正常供水和PEG-6000胁迫两种处理条件下D-应答蛋白组成一致，均由核酮糖-1,5-二磷酸羧化酶大亚基和磷酸甘油酸变位酶组成（P＜0. 05）；PEG-6000胁迫条件下两个蛋白（亚基）的鉴定得分分别为1 632和88，覆盖率分别为30%和21%；在鉴定的肽段中，有65个肽段同属于核酮糖-1,5-二磷酸羧化酶大亚基，其中35个肽段可信度均超过阈值分数；有5个肽段同属于磷酸甘油酸变位酶，其中4个肽段的可信度超过阈值分数。结果表明该蛋白条带可能主要由核酮糖-1,5-二磷酸羧化酶大亚基等组成。

英文摘要:

The expression of water stress responsive protein is significantly associated with drought resistance of wheat cultivars. This study aimed at understanding the type and characteristics of a water stress D-response protein. The seedling of winter wheat Jinmai 47 was treated under normal water supply and water stress using -0.5 MPa PEG-6000 solution for simulating drought treatments under controlled condition. The techniques of SDS-PAGE and NanoLC-MS/MS were applied to obtain the expression quantity and mass spectrometry identification of this D-response protein. SDS-PAGE results showed that the protein was expressed after treatment of 6 h, compared to normal water supply, and had the highest expression quantity under at the 48 h of -0.5 MPa PEG-6000 permeation stress treatment. Along with the extension of treatment, the expression of the protein gradually declined; when stressed for 48 h, the seedling was treated with normal water supply. The band of D-response protein was disappeared when rehydrated for 72 h. The target protein bands under normal water supply and water stress were excised from the gel and identified by mass spectrometry after the protein bands were digested by trypsin. Mass spectrometry identification obtained two positive peptides, and the results from two different treatment methods were identified. The results showed that the target protein bands were composed of ribulose -1,5-bisphosphate carboxylase (Rubisco) large subunit and phosphoglycerate mutase by mass spectrum analysis (P＜0. 05). The mass scores were 1 632 and 88 with coverage 30% and 21%, respectively. Among the matched peptides, there were 65 peptides that belong to ribulose-1, 5-bishosphate carboxylase (Rubisco) big subunit, among them,35 peptides's credibility scores were more than threshold credibility scores; There were five peptides which belong to phosphoglycerate mutase, and the credibility of four peptides of them were more than threshold credibility scores. The reasonable implications of the drought responsive protein were that it was Rubisco large subunit and other protein.

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