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作物学报 ›› 2014, Vol. 40 ›› Issue (07): 1190-1196.doi: 10.3724/SP.J.1006.2014.01190
王云鹏1,2,马景勇1,马瑞2,马建1,*,刘文国1,2,*
WANG Yun-Peng1,2,MA Jing-Yong1,MA Rui1,MA Jian 1,*,LIU Wen-Guo1,2,*
摘要：
EPSPS (5-烯醇丙酮莽草酸-3-磷酸合酶EC 2.5.1.19)是植物芳香族氨基酸和植物次生代谢产物生物合成中莽草酸途径的关键酶; 同时也是广谱性除草剂草甘膦的作用目标。本实验通过对草甘膦污染土壤宏基因组文库的建立及筛选, 成功克隆了一个新的草甘膦抗性的EPSPS基因(命名为soilEPSPS)。序列分析表明soilEPSPS基因全长1404 bp, 其编码的467个氨基酸中未涉及已公布专利中保护的氨基酸序列。原核功能验证表明该基因对草甘膦的耐受能力优于EPSPS CP4基因。将该基因与水稻Rubisco SSU引导肽相融合构建由actin启动子驱动的植物表达载体, 用农杆菌介导法实现了水稻的遗传转化。抗性再生植株的PCR和Southern杂交结果表明所获得的26株再生植株均为转基因阳性植株, 其中共有3个单拷贝转化事件。草甘膦抗性鉴定证明纯合体T2代植株能够耐受高达500 mmol L-1的草甘膦。本研究为转基因抗除草剂水稻新品种的培育奠定了基础。
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