小麦TaLhca基因的克隆及表达分析

为了从分子水分研究小麦的光合作用,该研究采用RT-PCR方法,从小麦品种‘百农207’的叶中克隆到1个捕光叶绿素a/b结合蛋白基因,命名为TaLhca。序列分析结果表明,TaLhca的编码区序列(coding DNA sequence,CDS)长810 bp,编码269个氨基酸,推测分子量为29.31 kD,等电点为8.69。TaLhca被定位于叶绿体,无信号肽,存在3个明显的跨膜区域,预测其蛋白结构含有典型的捕光叶绿素a/b结合蛋白功能域(chlorophyll a/b binding domain),为亲水性非分泌蛋白。蛋白序列比对和进化树分析表明,小麦与二穗短柄草(Brachypodium distachyon)和水稻(Oryza sativa)中的Lhca序列相似性最高,亲缘关系最近。启动子顺式作用元件预测表明,启动子区域包含多个光响应元件及逆境响应元件。实时荧光定量PCR分析表明,TaLhca基因在小麦根、茎、叶中均有表达。其中在叶中表达量最高,在根中表达量最低,且受NaCl、干旱、ABA、H2O2和低温胁迫表达增强,受黑暗胁迫表达降低。该研究结果为进一步解析小麦光合作用机理及其相关基因的诱导表达特性提供了依据。

Cloning and Expression Analysis of TaLhca Gene in Wheat

To study the molecular mechanism of photosynthesis,we cloned a wheat Lhca gene from the leaves of‘Bainong207’using RT-PCR,and was designated as TaLhca.The coding sequence(CDS)of this cloned Lhca gene is 810 bp in length,encoding a polypeptide of 269 amino acids.The deduced protein molecular weight was 29.31 kD and its theoretical isoelectric point was 8.69.Bioinformatics analysis showed that TaLhca is a hydrophilic non-secretory protein with three transmembrane domains and a chlorophyll a/b binding domain.Multiple sequence alignments and phytogenetic analysis of Lhca proteins showed TaLhca is most similar with Lhca from Brachypodium distachyon and Oryza sativa.Cis-regulatory element analysis revealed that light responsiveness elements,stress responsiveness elements are in the promoter region of TaLhca.The results of Real-time PCR analysis suggested that TaLhca expressed in root,stem and leaf,with the highest level in leaf,while the lowest level in root.Interestingly,the expression of TaLhca could be strongly induced by NaCl,drought,ABA and H2O2 and low temperature stress,while down regulated by darkness treatment.This results provided references for better understanding on the mechanism of photosynthesis and gene expression in wheat.