| 摘要: |
| 【目的】克隆小麦Mlo基因家族新成员，分析其编码蛋白的结构、进化地位以及其在小麦根、茎、叶组织和生物胁迫下的表达模式。【方法】采用电子克隆、RT-PCR技术，从小麦品种“水源11”中分离出1个Mlo的cDNA序列基因，对其进行克隆和序列分析；利用InterProScan、TMpred、SignalP等在线分析工具，预测TaMlo蛋白的保守结构域、跨膜结构、信号肽、细胞定位等特征；采用DNASTAR软件分别进行氨基酸序列同源性比对和进化树分析，并利用qRT-PCR技术对其表达谱进行分析。【结果】以玉米ZmMlo8基因为种子序列，克隆了小麦新基因TaMlo8（暂命名）cDNA序列，该序列长1 897 bp，ORF为1 497 bp，编码499个氨基酸；InterProScan和SignalP预测结果显示，其为Mlo家族成员，含信号肽和7个跨膜区；BLASTX结果显示，TaMlo8与玉米ZmMlo8蛋白的相似性达78%；系统进化树显示，TaMlo8与玉米ZmMlo8处于同一分支下，而与小麦其他Mlo家族成员处于不同的分支上，进一步说明TaMlo8为小麦Mlo家族的新成员；表达谱分析表明，TaMlo8是组成型低表达，在小麦根、茎、叶组织中的表达量基本一致，在小麦与条锈菌互作的非亲和体系中诱导表达，在小麦与白粉菌的感病反应中诱导表达。【结论】克隆到1个小麦Mlo基因新成员TaMlo8，其在小麦与条锈菌的抗性反应和小麦与白粉菌的感病反应中起一定作用，说明TaMlo8基因对条锈菌和白粉菌的抗病路径可能不同，为进一步研究其在小麦生物胁迫反应中的潜在功能奠定了基础。 |
| 关键词: 小麦条锈菌 小麦白粉菌 Mlo基因 电子克隆 结构域 |
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| 基金项目:国家自然科学基金项目（31071651）；现代农业产业技术体系建设专项（2006AA10A104）；高等学校学科创新引智计划项目（B07049） |
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| Cloning and expression analysis of a gene TaMlo8 in wheat |
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| Abstract: |
| 【Objective】The study was conducted to clone a gene of novel Mlo family members in wheat and to analyze the structure,evolution,function of the protein and expression profile.【Method】The electronic cloning technology and the RT-PCR were combined to obtain a new gene encoded Mlo from wheat Suwon 11 designated TaMlo8.Characteristics of the conserved domain,transmembrane structure,signal peptide,and localization of the encoded TaMlo protein were determined using bioinformatics analysis including InterProScan,TMpred and SignalP.The analysis of multiple alignment and phylogenetic tree was performed using DNASTAR software package.The expression patterns of the gene under biotic stresses were investigated by real time quantitative PCR.【Result】The wheat Mlo gene designated as TaMlo8 was isolated.The full-length of cDNA is 1 897 bp and the open reading frame (ORF) is 1 497 bp,encoding 499 amino acids.InterProScan demonstrated that its amino acid is the Mlo family member;The BLASTX result showed that TaMlo and the maize ZmMlo8 protein similarity reaches 78%;Phylogenetic trees showed TaMlo and maize ZmMlo8 is under the identical branch, but is in the different branch with the wheat other Mlo family members.The results further explained that TaMlo8 is the Mlo family’s another young member.The expression profiling of TaMlo8 was done,TaMlo8 gene presented almost the same expression in root,shoot and leaves.It is induced up expression in incompatible of wheat-Pst and in compatible of wheat-Bgt.【Conclusion】The TaMlo8 gene was cloned from wheat and characterized using a series of bioinformatics analysis.TaMlo8 gene is constitutionly low expression and involves resistance response in interaction of wheat-Pst and susceptible response in wheat-Bgt.It shows that TaMlo8 resistance pathway is different in two disease systems,which may lay a sound foundation for further functional analysis of the gene putatively involved in wheat. |
| Key words: Puccinia striiformis f.sp.tritici Blumeria graminis f.sp.tritici Mlo gene in silico cloning domain |
