论文部分内容阅读
多胺作为一类新型植物生理活性物质,对植物生长发育起着重要作用,在细胞工程与农业生产中具有广阔的应用价值。多胺氧化酶(Polyamine oxidase,PAO)是催化多胺降解的关键酶之一,对调控植物细胞中多胺浓度非常重要,影响植物的生长发育、形态建成过程。根据国际DNA数据库中已报道的PAO蛋白的保守结构域,设计简并引物(degenerate pri mer),通过RT-PCR方法首次从花生(仲恺花1号)中克隆PAO基因cDNA序列,并利用BLASTn、BLASTp及多重序列排比(Multiple Sequence Alignment)等生物信息学方法对所克隆的cDNA进行序列分析与鉴定,初步确定已从花生中克隆到PAO基因的同系物(homolog)。根据克隆的花生PAOcDNA序列(AhPAO1)设计特异引物,通过半定量RT-PCR方法检测花生胚根、胚芽和幼叶中AhPAO1基因的表达,结果发现胚根中AhPAO1基因表达最强,胚芽和幼叶中的表达无明显差异。本研究为进一步了解花生种子萌发和幼苗生长发育过程中多胺氧化酶基因表达的时空特异性奠定实验基础,并对进一步利用植物基因工程技术综合调控花生体内多胺水平有重要实践意义。
As a new type of plant physiologically active substances, polyamines play an important role in plant growth and development and have broad application value in cell engineering and agricultural production. Polyamine oxidase (PAO) is one of the key enzymes that catalyze the degradation of polyamines. It is very important to regulate the concentration of polyamines in plant cells, affecting the growth and morphogenesis of plants. According to the conserved domain of PAO protein reported in the international DNA database, a degenerate primer was designed. The cDNA sequence of PAO gene was cloned from peanut (Zhongkaihua No.1) by RT-PCR. BLASTn, BLASTp and Multiple Sequence Alignment were used to sequence and identify the cloned cDNA. The homolog of PAO gene was cloned from peanut. Specific primers were designed according to the cloned peanut PAOcDNA sequence (AhPAO1). The expression of AhPAO1 gene in radicle, germ and young leaves of peanut was detected by semi-quantitative RT-PCR. The results showed that the expression of AhPAO1 was the highest in radicle, In the expression of no significant difference. This study laid the experimental foundation for further understanding the temporal and spatial specificity of polyamine oxidase gene expression in peanut seed germination and seedling growth and development and has important practical significance for the further utilization of plant genetic engineering technology to comprehensively regulate polyamine levels in peanut.