水稻OsRZFP34基因逆境表达特征及其启动子的克隆与分析

Expression patterns of OsRZFP34 under diverse stress treatments and its promoter’s cloning and characterization

采用qRT–PCR技术分析水稻OsRZFP34在高温(42℃)、低温(4℃)、10%聚乙二醇(PEG)6000模拟干旱、高盐(100 mmol/L NaCl)和脱落酸(ABA)处理(100μmol/L)下的表达模式;克隆OsRZFP34基因启动子,并利用在线软件New PLACE和PlantCARE对其序列进行顺式作用元件分析;构建OsRZFP34pro::GUS表达载体并转化水稻;通过组织化学染色和GUS酶活性检测,分析在不同胁迫处理条件下OsRZFP34启动子驱动GUS基因表达的活性。qRT–PCR分析结果表明,上述5种处理均能不同程度地诱导OsRZFP34的表达,其中高温胁迫对其表达的诱导程度最强,而ABA对其表达只有微弱的诱导。顺式作用元件分析结果表明,该启动子上包含有多个与逆境响应、激素响应以及Ca^2+信号转导相关元件。GUS组织化学染色和荧光分析结果显示,OsRZFP34基因启动子的活性受到高温、低温、PEG、盐和ABA的调控,表明该启动子是受多种逆境诱导的诱导型启动子。

In this study,qRT-PCR was used to analyze the expression patterns of OsRZFP34 under various conditions including the high temperature(42℃),low temperature(4℃),simulated drought with 10%polyethylene glycol(PEG)6000,salt(100 mmol/L NaCl)and ABA(100μmol/L)treatments.The promoter of OsRZFP34 was cloned and its sequence was analyzed using online software New PLACE and PlantCARE;The OsRZFP34pro::GUS expression vector was constructed and transformed into rice;The activities of OsRZFP34 promoter driving GUS gene expression under different stress conditions were analyzed by histochemical assay and GUS enzyme detection.The results of qRT-PCR showed that the expression of OsRZFP34 could be induced by the above treatments.High temperature stress had the strongest induction on the expression of OsRZFP34,while ABA had only a weak induction on its expression.The results of the cis-regulatory element analysis show that,the promoter sequence contains a number of elements related to stress response,hormone response and Ca^2+signal transduction.The results of histochemical assay and quantitative GUS fluorescence analysis showed that the activity of OsRZFP34 promoter was regulated by high temperature,low temperature,PEG,salt and ABA treatments,which indicated that OsRZFP34 promoter was a multi-stress-inducible promoter.