苎麻atp6和atp9基因的克隆表达及与细胞质雄性不育的相关性.pdfVIP

HEREDITAS (Beijing) 2008 11 , 30(11): 1487―1498 ISSN 0253-9772 研究报告 DOI: 10.3724/SP.J.1005.2008.01487 atp6 atp9 段继强, 杜光辉, 李建永, 梁雪妮, 刘飞虎 , 650091 : 根据GenBank 报道的双子叶植物线粒体atp6 和atp9 基因编码区保守序列设计简并引物, 通过PCR 技术 从苎麻细胞质雄性不育系、保持系和恢复系(简称“ 三系”) mtDNA 中扩增目的基因片段, 发现所得序列开放阅读 框虽不完整, 但与 GenBank 报道的其他植物线粒体atp6 和atp9 基因同源性分别高于 94%和 85%。采用DNA Walking 步移法分别从 3 端和 5 端扩增两个基因片段的未知侧翼序列, 分离出完整的苎麻线粒体 atp6 和 atp9 基因, 包含了完整的开放阅读框。其中“ 三系” 的atp6 基因在mtDNA 水平、转录和翻译调控水平、蛋白质水平 上均无差异。不育系atp9 基因在编码区3 端与保持系和恢复系相比存在若干个碱基的差异和缺失; RT-PCR 分 析还表明, 不育系atp9 基因在现蕾期和盛花期的表达量很高。推测不育系atp9 基因的结构变异和/或异常表达 与苎麻细胞质雄性不育(CMS) 的关系密切。 : 苎麻; 细胞质雄性不育; atp6 基因; atp9 基因; DNA Walking; RT-PCR Cloning and expression of atp6 and atp9 genes from ramie (Boehmeria nivea (L.) Gaud.) and their relationship with cytoplasmic male sterility DUAN Ji-Qiang, DU Guang-Hui, LI Jian-Yong, LIANG Xue-Ni, LIU Fei-Hu Laboratory of Plant Improvement and Utilization , Yunnan University, Kunming 650091, China Abstract: The atp6 and apt9 gene fragments associated with cytoplasmic male sterility (CMS) were cloned from the mito- chondrial DNA of a ramie (Boehmeria nivea (L.) Gaud.) cytoplasmic male sterile line and its maintainer and restorer lines using PCR and degenerated primer strategy. The primers were designed according to the reserved sequences in the encoding region of mitochondrial genes atp6 and atp9 of some dicotyledons from GenBank. These fragments did not have complete encoding region but showed the homology of 94% and 85% with atp6 and atp9 genes from the referred dicotyledons in GenBank. The complete atp6 and atp9 genes including the complete open reading frames were cloned by means of ampli- fying the 3′ and 5′end unknown sequences of thes