棉花PGHNBS启动子维管束特异表达元件的分离及功能研究

Isolation and functional analysis of the vascular bundle specific element of promoter PGHNBS in cotton

PGHNBS是来源于陆地棉的一个CC-NBS-LRR类基因GHNBS的启动子,该启动子具有维管束特异表达特征。为了分离PGHNBS启动子的调控元件,对该启动子进行了缺失分析。将PGHNBS启动子6个部分缺失片段分别与GUS基因融合构建植物表达载体,这6个表达载体分别为PGN15(-1 420 bp～-28 bp)、PGN27(-1 300 bp～-28 bp)、PGN31(-1 071 bp～-28 bp)、PGN44(-959 bp～-28 bp)、PGN53(-807 bp～-28 bp)和PGN996(-604 bp～-28 bp)。用植物表达载体转化拟南芥,通过对转基因植物的GUS组织化学染色,进一步确认了韧皮部特异表达元件位于-1 071 bp～-959 bp区域。用PLACE进行相似性比较,发现该区域含有脱落酸(ABA)诱导元件、铜离子响应元件以及多个与抗逆相关的MYB转录因子的识别位点。另外,在-1 559 bp～-1 420 bp区域中存在增强子元件。

PGHNBS is the promoter of a CC-NBS-LRR gene GHNBS of cotton,which owns the vascular bundle specific character and drives the objective gene expressing in the vascular bundle.To isolate the vascular bundle specific element of PGHNBS,deletion analysis was conducted.Six varied length promoter sequences were fused to the GUS gene separately to construct the plant expression vector.The six vectors were PGN15（-1 420 bp——28 bp）,PGN27（-1 300 bp——28 bp）,PGN31（-1 071 bp——28 bp）,PGN44（-959 bp——28 bp）,PGN53（-807 bp——28 bp） and PGN996（-604 bp——28 bp）,which were transformed to the Arabidopsis afterwards.The GUS staining of transformed plants showed the vascular bundle specific element was located within the fragment of-1 071 bp——959 bp.Similarity comparison by PLACE revealed abscisic acid（ABA） inductive element,copper ion responsive element and some stress resistance related MYB transcription factor recognition locus were located within this region.In addition,some enhancer elements was located within the region of-1 559 bp——1 420 bp.