香蕉凝集素基因启动子的分离、序列分析及鉴定

香蕉是世界上最重要的水果之一,由于香蕉果实是利用转基因方法生产重组药用蛋白或有价值的化合物的理想器官,构建能在香蕉果实中高水平表达异源蛋白质的表达载体是非常有意义的。而一个高效表达的载体,启动子则是其最重要元件之一,因此,果实特异性表达启动子的获得是香蕉作为生物反应器的前提。香蕉凝集素是一种在香蕉果实中大量存在的蛋白质,其基因被证明在果肉组织中大量表达。利用染色体步移法克隆到香蕉凝集素基因5′端上游的一段长702bp的序列,经序列测定及软件分析表明,该序列具有典型的启动子结构。此序列置换植物表达载体pBI121的CaMV35S启动子,构建植物表达载体,命名为pBIL2,该启动子下游为gus基因。利用基因枪法转化香蕉的根、叶和果实薄片,对gus基因的瞬时表达进行测定,结果表明所获得的凝集素基因启动子,只在香蕉果肉中瞬时表达,该启动子的表达具有果实特异性,并且表达量较高。

Isolation, Sequencing Analysis and Characterization of the Promoter of Banana Lectin Gene

Banana (Musa spp) is one of the most important fruit crops in the world. Banana fruit is an ideal organ for producing foreign pharmaceutical proteins and chemicals by genetic engineering. A perfect promoter driving foreign gene to express strongly and specifically in banana fruit is necessary for that. In order to isolate a banana fruit-specific expressed promoter, a fragment of 702 nt nucleotide sequence upstream 5' of banana lectin (BanLec) gene, which was demonstrated to express specifically in banana fruit previously, was isolated by using chromosomal walking in this study. Bioinformatical analysis of this sequence shows that the sequence contains some typical elements of a promoter. To identify the fruit-specific expression of this promoter, a construct was derived from pBI121, which originally CaMV 35S promoter was replaced by the 702 nt nucleotide sequence, and named as pBIL2. Transformations of pBIL2 to roots, leaves and fruit pieces of banana were carried out by using particle bombardment. The transient expression of gus showed that the gus expressed specifically in banana fruit with a little higher level compared with CaMV 35S. It is the first report that BanLec promoter is a potential fruit-specific expressed promoter which can further be used in transgenes into banana.