不结球白菜过氧化还原蛋白基因Brc2-Cys Prx的克隆和表达分析

发布时间：2018-03-24 14:57

本文选题：不结球白菜　切入点：霜霉病菌　出处：《南京农业大学学报》2017年01期

【摘要】：[目的]双半胱氨酸型硫氧还蛋白过氧化物酶Brc2-Cys Prx是过氧化还原蛋白Peroxiredoxins(Prxs)的亚家族。Prxs是植物抗氧化酶防御系统的成员,对清除植物体内活性氧具有重要作用。本文旨在克隆和研究Brc2-Cys Prx基因在不结球白菜处于不同胁迫条件下的表达模式。[方法]以不结球白菜霜霉病抗病自交系‘苏州青’和霜霉病感病自交系‘矮脚黄’为试验材料,利用RACE技术克隆得到不结球白菜Brc2-Cys Prx基因。利用生物信息学方法对该基因进行信息学分析。采用qRT-PCR技术对不结球白菜叶片Brc2-Cys Prx基因在霜霉病菌侵染及NaCl、CdCl_2、PEG6000和ABA等非生物胁迫处理条件下的表达模式进行研究。[结果]Brc2-Cys Prx基因的c DNA全长为999 bp,其中开放阅读框长度为810 bp,共编码270个氨基酸。该蛋白不存在信号肽序列,预测相对分子质量为28.15×10~3,理论等电点为5.87。不结球白菜Brc2-Cys Prx蛋白的二级结构主要由α-螺旋和无规则卷曲构成。该蛋白含有1个PRX_Typ2cys结构域,属于硫氧还蛋白超家族。系统进化分析表明:不结球白菜Brc2-Cys Prx蛋白与同属植物的进化关系相近,其中与大白菜和甘蓝型油菜进化关系最近。霜霉病菌侵染条件下,Brc2-Cys Prx基因表达呈逐渐上调趋势,在‘矮脚黄’中的表达与‘苏州青’相比上调迟钝,且该基因在‘矮脚黄’中的表达量一直低于‘苏州青’。0.2 mol·L~(-1)NaCl和200 g·L~(-1)PEG6000处理抑制Brc2-Cys Prx基因的表达;150μmol·L~(-1)CdCl_2处理促进Brc2-Cys Prx基因的表达,在24 h达到最大值;100μmol·L~(-1)ABA处理后,Brc2-Cys Prx基因表达上调,而12 h后,表达受抑制。[结论]霜霉病菌侵染条件下,Brc2-Cys Prx在抗性系‘苏州青’和感病系‘矮脚黄’中的表达存在差异,并且在抗病系中的表达量较高,推测Brc2-Cys Prx基因的高转录水平是抗病品种较之感病品种对霜霉病菌侵染具有更高抗性的原因之一。在非生物胁迫条件下,Brc2-Cys Prx基因具有不同的应答模式,推测可能是涉及到不同的物质或信号途径的原因。
[Abstract]:[objective] the subfamily of biscysteine thioredoxins peroxidase (Brc2-Cys Prx) is a member of the antioxidant enzyme defense system of plants. The purpose of this paper is to clone and study the expression pattern of Brc2-Cys Prx gene in non-heading Chinese cabbage under different stress conditions. [methods] the resistant inbred line'Su'of non-heading cabbage downy mildew was used. Zhou Qing 'and downy mildew susceptible inbred line' Dailihuang 'were used as experimental materials. The Brc2-Cys Prx gene of non-heading cabbage was cloned by RACE technique. The gene was analyzed by bioinformatics. The Brc2-Cys Prx gene of non-heading cabbage leaves was infected by qRT-PCR technique in downy mildew and non-#en6# and NaCl-CdCl2PEG6000, etc. The expression pattern of Brc2-Cys Prx gene was studied under biological stress. [results] the total length of c DNA of Brc2-Cys Prx gene was 999bp, and the length of open reading frame was 810 BP, encoding 270 amino acids. The predicted molecular weight is 28.15 脳 10 ~ (-3) and the theoretical isoelectric point is 5.87. The secondary structure of Brc2-Cys Prx protein in non-heading Chinese cabbage is mainly composed of 伪 -helix and irregular curl. The protein contains a PRX_Typ2cys domain. Phylogenetic analysis showed that Brc2-Cys Prx protein in non-heading Chinese cabbage was similar to that in the same genus. Under the condition of downy mildew infection, the expression of Brc2-Cys Prx gene was gradually up-regulated, and the expression of Brc2-Cys Prx in 'low foot yellow' was slowly up-regulated compared with 'Suzhou Qinghuang', and was closely related to the evolution of Brc2-Cys in Brassica campestris and Brassica napus under the condition of downy mildew. Moreover, the expression of this gene was significantly lower than that of 'Suzhou Qingyao. 0.2 mol L~(-1)NaCl and 200g L~(-1)PEG6000 treatment, which inhibited the expression of Brc2-Cys Prx gene (150 渭 mol L~(-1)CdCl_2) and increased the expression of Brc2-Cys Prx gene. The expression of Brc2-Cys Prx gene was up-regulated after treatment with 100 渭 mol L~(-1)ABA at 24 h after treatment with 100 渭 mol L~(-1)ABA. After 12 h, the expression of Brc2-Cys Prx was inhibited by downy mildew. [conclusion] the expression of Brc2-Cys Prx in the resistant lines' Suzhou green 'and the susceptible line' Daijiihuang 'was different, and the expression of Brc2-Cys Prx was higher in the resistant lines. It is inferred that the high transcription level of Brc2-Cys Prx gene is one of the reasons why resistant varieties have higher resistance to downy mildew than susceptible varieties. The Brc2-Cys Prx gene has different response patterns under abiotic stress. Speculation may be involved in different substances or signaling pathways.
【作者单位】：南京农业大学作物遗传与种质创新国家重点实验室/农业部华东地区园艺作物生物学与种质创新重点实验室/园艺学院;
【基金】：江苏省农业科技自主创新项目[CX(15)1015] 江苏省科技支撑计划项目(BE2013429,BE2012325) 国家自然科学基金项目(31272173)
【分类号】：S634.3

【相似文献】