紫花苜蓿MsLEA4启动子的克隆及表达分析

为了深入研究紫花苜蓿（Medicago sativa）MsLEA4基因的功能，本试验采用染色体步移技术从紫花苜蓿基因组中扩增出MsLEA4启动子序列，构建GUS植物超表达载体，并根据序列分析结果对紫花苜蓿进行相应逆境胁迫。结果表明：紫花苜蓿MsLEA4启动子含有响应脱落酸（abscisic acid，ABA）调控的顺式作用元件ABRE，响应赤霉素（gibberellin，GA）调控的顺式作用元件P-box，参与胚乳合成的顺式作用元件GCN4和Skn-1，以及涉及光调控和光周期的顺式作用元件；外源ABA、GA、持续光照以及黑暗均能诱导MsLEA4基因的表达；GUS基因只在拟南芥花和荚果中表达。因此，MsLEA4基因能参与紫花苜蓿的逆境调控，与该基因启动子序列中所含的一系列顺式作用元件相关。本研究为进一步探索MsLEA4基因在紫花苜蓿逆境胁迫、光调控以及组织特异性表达中的作用奠定了基础。

Cloning and Expression Analysis of MsLEA4 Promoter from Medicago Sativa

In order to further analyze the function of MsLEA4,we cloned MsLEA4 promoter sequence from alfalfa (Medicago Sativa) genome by genomic walking technique,constructed MsLEA4 promoter::GUS transient expression vector,and also conducted corresponding stresses on alfalfa according to the promoter sequence analysis. The results showed that: MsLEA4 promoter contained the basic elements TATA-box and CAAT-box,included abscisic acid (ABA) responsive elements (ARBE),gibberellin (GA) responsive elements (P-box),and cis-elements (GCN4,Skn-1) involved in endosperm synthesis. In addition to this,the promoter sequence also contained 6 cis-elements related to light regulation and a cis-element related to circadian rhythm. ABA,GA,continuous light and darkness all can induce MsLEA4 expression; the GUS gene was expressed only in flowers and siliques of Arabidopsis thaliana. Therefore,MsLEA4 can participate in the regulation of alfalfa stress responses,which is related to a series of cis-elements contained in the promoter. This study laid a foundation for further exploring the role of MsLEA4 in stresses,light regulation and tissue specific expression of alfalfa.