六倍体大小麦tritordeum花药组织特异性启动子的鉴定与分离

启动子是基因表达调控的重要顺式元件,也是基因工程表达载体的一个重要元件。一个无启动子的带有UidA基因的质粒pPLGUS通过基因枪转化进tritordeum材料中,对转基因材料的多种不同组织进行了X-gluc显色来检测不同组织中的GUS活性,有一个株系的花药组织特异性启动子已被证明成功捕获,并通过PCR方法将其分离。提取叶片的总DNA作模板,上游使用水稻花药启动子分离的引物P1,以UidA基因的部分序列为下游引物P2,PCR扩增UidA基因的上游旁侧序列。已经获得一条长667 bp的目的片断,含有部分UidA基因的序列和一段UidA基因的上游旁侧序列,该序列中具有植物启动子的一些必备元件,初步断定它是一段花药组织特异性启动子序列。

Tagging and Isolation of the Anther-specific Promoter in Hexaploid Tritordeum

The promoter is an important element of gene expression,and tissue specific promoters are very useful in gene engineering.A promoterless UidA containing plasmid derived from pPLGUS was transformed into tritordeum.Histochemical analysis of various tissues in transgenic tritordeum was carried to examine tissue specific expression of GUS activity.A transformant line with anther-specific GUS expression was successfully labeled and isolated.PCR method was used to isolate this anther-specific promoter with DNA extracted from leave as template,using primer pair of rice anther specific promoter,primer P1 as forward primer,UidA gene specific sequence as reverse primer.By sequencing and analyzing the amplified DNA fragment from PCR reaction,a part of UidA gene and a flanking sequence were obtained.Some essential promoter elements were found in this sequence.Therefore the fragment obtained was believed to be a part of anther specific promoter.