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小麦再生相关基因 TaDCC1-2B的克隆与功能验证
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引用本文:贾冬冬,王京宏,崔桂宾,张舒梦,张 超,孙风丽,王竹林,奚亚军.小麦再生相关基因 TaDCC1-2B的克隆与功能验证[J].西北农业学报,2019,28(12):1948~1959
DOI:10.7606/j.issn.1004-1389.2019.12.006
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作者单位
贾冬冬,王京宏,崔桂宾,张舒梦,张 超,孙风丽,王竹林,奚亚军 (1. 西北农林科技大学 农学院, 陕西杨凌 7121002. 农业农村部西北地区小麦生物学与遗传育种重点实验室, 陕西杨凌 712100) 
基金项目:陕西省种质资源保护与利用(20171010000004);杨凌示范区农业科技推广项目(2017-TS-20)。
中文摘要:硫氧还蛋白DCC1在拟南芥中依赖氧化还原修饰通过调节活性氧(ROS)的稳态来调控植物的再生。为深入了解小麦中该基因的序列特征及功能,从普通小麦‘中国春’中克隆获得一个小麦硫氧还蛋白基因 TaDCC1-2B,其CDS长度为645 bp,编码214个氨基酸。生物信息学分析表明,该基因编码的蛋白分子质量为23.59 ku,理论等电点为8.94,亲水性总平均值为-0.409,推测为亲水性蛋白。系统进化分析表明该基因在不同物种间具有较强的保守性,启动子区域含有与分生组织形成和赤霉素响应相关的的顺式作用元件。利用农杆菌介导法将 TaDCC1-2B 基因转入拟南芥硫氧还蛋白突变体dcc1中,转基因植株根尖诱导的再生芽的再生率接近于野生型,其体内 TaDCC1-2BWUS基因的表达水平也显著高于突变体,表明转基因株系中 TaDCC1-2B基因的过量表达维持了愈伤组织中内源ROS水平的稳定,使得WUS基因得以正常表达,从而维持了拟南芥愈伤组织的再生能力,推测 TaDCC1-2B基因可能在小麦组织培养过程中也将发挥相似作用。
中文关键词:小麦(Triticum aestivum L.)  硫氧还蛋白   TaDCC1-2B基因  ROS内稳态  体外再生
 
Cloning and Functional Verification of Regeneration Related Gene TaDCC1-2B in Wheat
Abstract:Thioredoxin DCC1 in Arabidopsis thaliana depends on redox modification to regulate plant regeneration by regulating the homeostasis of reactive oxygen species (ROS).In order to further understand the sequence characteristics and function of the gene in wheat, a wheat thioredoxin gene TaDCC1-2B, was cloned from common wheat ‘Chinese Spring’,whose CDS with a length of 645 bp, encoding 214 amino acids.Bioinformatics analysis showed that the molecular mass of the protein encoded by the gene is 23.59 ku.The theoretical isoelectric point is 8.94,and the total average hydrophilicity is -0.409,which was presumed to be a hydrophilic protein.Phylogenetic analysis showed that the gene is highly conserved among different species.The promoter region contains cis-acting elements related to meristem formation and gibberellin response. TaDCC1-2B gene was transferred into Arabidopsis thaliana thioredoxin mutant dcc1 by Agrobacterium tumefaciens-mediated method, the regeneration rate of regenerated buds induced from root tip of transgenic plants was close to that of wild type, and the expression levels of TaDCC1-2B gene and WUS gene in transgenic plants were significantly higher than those in mutants.The results showed that the overexpression of TaDCC1-2B gene in transgenic lines maintained the stability of endogenous ROS level in callus and enabled the normal expression of WUS gene,thus maintaining the regeneration ability of Arabidopsis thaliana callus. It is speculated that TaDCC1-2B gene may play a similar role in wheat tissue culture.
keywords:Wheat(Triticum aestivum L.)  Thioredoxin   TaDCC1-2B gene  ROS homeostasis  Invitro regeneration
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