The construction of plasmids suitable for searching important rice genes.

适合寻找重要水稻基因的质粒的构建。

• 批准号: 03556002
• 负责人: HIRAI Atsushi
• 金额: $ 10.82万
• 依托单位: The University of Tokyo (1992-1993)Nagoya University (1991)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B)
• 财政年份: 1991
• 资助国家: 日本
• 起止时间: 1991 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-03556002/
• 关键词: rice; nuclear DNA; particle gun; mutation; mitochondrial DNA; nad9; nad9; 巨大DNA; パ-ティクルガン

In order to clone unknown but agriculturally useful genes, we have constructed a plasmid suitable for transferred-DNA-mediated insertional mutagenesis. It has a 35S promoter and a chloramphenicol acetyl transferase gene which are active in plant cells, and a replication origin and a ampicillin resistant gene which are active in bacterial cells. We tried to transform this 7.6 kb long plasmid into rice culture cells by a particle gun, but we could not obtain transformants. The mitochondrial genomes of higher plants are much larger and more complex than those of other eukaryotic organisms. However, number of identified genes are limited. We speculated that mitochondrial DNA codes more active genes and searched unidentified important genes in rice mitochondrial DNA.First, we constructed a transcription map of the mitochondrial genome from rice. Then we determined nucleotide sequence the area in where the RNA is transcribed but genes are not identified. We found nad9, the gene for NADH dehydrogenase subunit 9, by this method.

为了克隆未知但在农业上有用的基因，我们构建了一种适合于转移DNA介导的插入突变的质粒。它具有在植物细胞中活跃的35S启动子和氯霉素乙酰转移酶基因，以及在细菌细胞中活跃的复制起点和氨苄西林抗性基因。我们试图用粒子枪将这个7.6kb长的质粒转化到水稻培养细胞中，但未能获得转化子。高等植物的线粒体基因组比其他真核生物的要大得多，也更复杂。然而，已识别的基因数量有限。我们推测线粒体DNA编码更多的活性基因，并在水稻线粒体DNA中寻找未知的重要基因。首先，我们构建了水稻线粒体基因组的转录图谱。然后我们确定了核苷酸序列，这是RNA被转录但基因未被识别的区域。通过这种方法，我们发现了NADH脱氢酶亚单位9的基因NAD9。

项目成果

Kanno,A.and A.Hirai: "Comparative studies on the structure of chloroplast DNA from four species of Oryza:Cloning and physical map." Theor.Appl.Genet.

Kanno,A. 和 A.Hirai：“四种稻叶绿体 DNA 结构的比较研究：克隆和物理图谱”。

Kanazawa,A.: "Restriction fragments homologous to mitochondrial plasmid-like DNAs are located within limited chromosomal regions on the rice nuclear genome." Theor.Appl.Genet.87. 577-586 (1993)

Kanazawa,A.：“与线粒体类质粒 DNA 同源的限制性片段位于水稻核基因组的有限染色体区域内。”

Honda,H.and A.Hirai: "The gene for the alpha-subunit of ATPase:a site of homologous recombination in plant mitochondrial DNA also functions in somatic hybrid cells." Theor,Appl.Genet.

Honda，H. 和 A.Hirai：“ATP 酶 α 亚基的基因：植物线粒体 DNA 中的同源重组位点，也在体细胞杂交细胞中发挥作用。”

Kanno,A.: "Variations in chloroplast DNA from rice(Oryza sativa)differences between deletions mediated by short direct-repeat sequences within a single species." Theor.Appl.Genet.86. 579-584 (1993)

Kanno,A.：“水稻 (Oryza sativa) 叶绿体 DNA 的变异与单个物种内短直接重复序列介导的缺失之间存在差异。”

Hatano,S.: "The preparation of high-molecular-weight DNA from rice and its analysis by pulsed-field gel.electrophoresis." Plant Science. 83. 55-64 (1992)

Hatano,S.：“从水稻中制备高分子量 DNA 及其脉冲场凝胶电泳分析。”