【作者】 徐云峰；

【导师】 李德全；

【作者基本信息】 山东农业大学 ， 植物学， 2005， 硕士

【摘要】 本文以抗逆（旱）性较强的玉米品种LD9002 为试验材料,采取溶液培养幼苗,信号调节物质、低温和干旱胁迫的处理方式,对玉米根系中的蛋白磷酸酶基因进行了分离克隆、表达特性及转基因植株生理功能的研究。运用RT-PCR 方法,首次从玉米根系中分离到一个PP2C 基因的cDNA克隆, 通过Southern 杂交、Northern 杂交、转化等技术对该基因的组成情况、表达特性和生理功能进行了初步研究,旨在探讨玉米等禾谷类作物抗逆的分子机制,为改良农作物抗逆性提供理论依据,主要结果如下: 1. 依据植物蛋白磷酸酶2C（protein phosphatase 2C,PP2C）基因的保守区设计简并引物,运用RT-PCR 方法,从玉米根系中分离出一个长度为936bp 的PP2C 基因的cDNA 克隆, 命名为ZmPP2C。GenBank注册号为AY621066。2. 提取正常生长状态下的玉米基因组DNA,用BamHI,EcoRV,SacI 三种限制性内切酶对玉米基因组DNA 进行完全酶切,以ZmPP2C 基因5′端为模板合成探针进行Southern 杂交。结果表明:ZmPP2C 基因在玉米基因组中可能是两个拷贝,弱带可能是其他同源基因,在玉米中存在一个小的PP2C 基因家族。3. 提取玉米根茎叶的总RNA,以α-³²P-dCTP 标记的ZmPP2C 基因5′端为探针进行Northern 杂交,结果表明,不同玉米组织中ZmPP2C 基因的表达差异明显。用CaCl2 、ABA、MgCl₂ 、EGTA、PEG 处理玉米根系24 h 后,对ZmPP2C 在玉米根系中的表达进行Northern 杂交分析,更多还原

【Abstract】 The maize variety of stress-resistance LD9002 was cultivated by solution in the mode of sitmulated treatments with signalling regulators, low-temperature and drought stress. A homologous gene of protein phosphatase 2C was cloned from Zea mays roots for the first time, the state of the gene in genome, the expression characteristics, the physiology functions were studied in this thesis by Southern blot, Northern blot and transforming technology and so on. Researching is to explore the molecular mechanisms of stress-resistant of maize and other crops, and to privide the basis for the screening and utilizing of stress-tolerance resource. The main results were as follows: 1. According to the conserved motifs of plant protein phosphatase 2C gene, degenerate oligonucleotides were designed. A full cDNA sequence of PP2C gene from Zea mays roots was cloned by RT-PCR. It was named ZmPP2C and had 936 bp . The accession number is AY621066 in GenBank. 2. Genomic DNA was isolated from normal growth maize, and completely digested with three restriction endonucleases: BamH I, EcoR V, Sac I. Using N-sequence of ZmPP2C gene as probe, southern blot showed that the ZmPP2C gene was a low copy in the Zea mays genome, and there was probably a small PP2C gene family. 3. Total RNA was isolated form root, stem and leaf in maize. Using N-sequence of ZmPP2C gene as probe, Northern blot showed that the更多还原