玉米醇溶蛋白ze19基因启动子克隆与功能分析

Cloning and Functional Analysis of a Zein ze19 Promoter

从玉米自交系9801基因组DNA中克隆玉米醇溶蛋白基因ze19启动子,将其克隆到pMD18-T载体上。该序列与发表序列同源性为94%,包括TATAbox、CAATbox启动子基本元件以及多种参与调节醇溶蛋白表达的顺式元件。将ze19启动子取代质粒PBI121-gus中的CaMV35S启动子,构建由ze19启动子驱动GUS的植物表达载体pBIze19-gus,利用土壤杆菌介导法将重组载体pBIze19-gus转入烟草中。对转基因烟草植株GUS活性的定性与定量分析结果表明,ze19启动子驱动GUS基因在转基因烟草种子中表达活性最高,在叶片中活性很弱,在茎中没有活性。所克隆的ze19启动子具有种子特异表达特性,为玉米种子生物反应器的研究提供借鉴。

The zein zel9 promoter was amplified using genomic DNA of maize inbred line 9801, and the se- quence was cloned to vector pMD 18-T. Sequence analysis indicated that the cloned sequence shares 94% homologues with the published sequence, the cloned promoter contains the two basic promoter element, TATA box and CAAT box and several c/s-regulatory elements that necessary to mediate endosperm expression of zein. The plant expression vector pBIzel9-gus was constructed by replacing the CaMV35S promoter in plasmid PBI121-gus and placed up- stream of the ^-glueuronidase（GUS） reporter gene, tobacco plants were transformed via Agrobacterium tumefacients mediated procedure. The results of both GUS histoehemieal staining and fluorometrie quantitative analysis of GUS ac- tivity showed that the expression level of GUS fusion gene was significantly stronger in seed, lower in leaf and no ac- tivity in stem tissue. The results showed that the cloned promoter was seed specific in initiating target gene expression, which provides a useful tool for seed bioreaetor development.