诸葛菜EPSPS基因5′端的克隆和序列分析

Cloning and Sequencing the 5′ Fragments of EPSPS Gene from Orychophragmus violaceus

烯醇式丙酮基莽草酸 3 磷酸合成酶(5 enolpyruvylshikimate 3 phosphatesynthase,EP SPS)是植物和微生物莽草酸途径中的一个必需合成酶,植物中该基因的超量表达或该基因中某些氨基酸突变可以产生对除草剂草甘膦(glyphosate)的耐性.采用5′ RACE技术从诸葛菜(Orychophragmusviolaceus)叶中克隆出一条长743bp的cDNA片段.序列分析结果表明:该cDNA与其它植物的EPSPS基因具有相当高的核苷酸序列同源性.其所编码的氨基酸序列与欧洲油菜(Brassicanapus)同源性最高为90%,与拟南芥(Arabidopsisthaliana)、玉米(Zeamays)、西红柿(Lycopersiconesculentum)、烟草(Nicotianatabacum)和水稻(Oryzasativa)的同源性分别为88%,81%,64%,62%和71%.

5enolpyruvylshikimate3phosphate synthase is a necessary synthase for plants and microbes among the shikimate pathway. Plants can obtain glyphosatetolerance if the enzyme of this gene was overproduced and accumulated or a herbicideinsensitive enzyme produced due to some amino acid mutation. A cDNA fragments was isolated from Orychophragmus violaceus leaf using RTPCR and 5′RACE technique. The cDNA fragment is 743bp long, which can encode 219 amino acids. The nucleotide sequence analysis showed high similarity to other plants. The deduced protein sequence analysis showed 90% similarity to Brassica napus, 88% to Arabidopsis thaliana, 81% to Zea mays, 64% to Lycopersicon esculentum, 62% to Nicotiana tabacum and 71% to Oryza sativa.