Ito Takuya, Motohashi Reiko, Kuromori Takashi, Mizukado Saho, Sakurai Tetsuya, Kanahara Hiroko, Seki Motoaki, Shinozaki Kazuo
Laboratory of Plant Molecular Biology, Plant Functional Genomics Research Group, Genomic Sciences Center, RIKEN, 3-1-1 Koyadai, Tsukuba 305-0074, Japan.
Plant Physiol. 2002 Aug;129(4):1695-9. doi: 10.1104/pp.002774.
We transposed Dissociation (Ds) elements from three start loci on chromosome 5 in Arabidopsis (Nossen ecotype) by using a local transposition system. We determined partial genomic sequences flanking the Ds elements and mapped the elements' insertion sites in 1,173 transposed lines by comparison with the published genomic sequence. Most of the lines contained a single copy of the Ds element. One-half of the lines contained Ds on chromosome 5; in particular, insertion "hot spots" near the three start loci were clearly observed. In the other lines, the Ds elements were transposed across chromosomes. We found other insertion hot spots at the tops of chromosomes 2 and 4, near nucleolus organizer regions 2 and 4, respectively. Another characteristic feature was that the Ds elements tended to transpose near the chromosome ends and rarely transposed near centromeres. The distribution patterns differed among the three start loci, even though they possessed the same Ds construct. More than one-half of the Ds elements were inserted irregularly into the genome; that is, they did not retain the perfect inverted repeat sequence of Ds nor leave perfect target site duplications. This precise analysis of distribution patterns will contribute to a comprehensive understanding of the transposing mechanism. From these Ds insertion sites, we have constructed a database for screening gene-knockout mutants in silico. In 583 of the 1,173 lines, the Ds elements were inserted into protein-coding genes, which suggests that these lines are gene-knockout mutants. The database and individual lines will be available freely for academic use from the RIKEN Bio-Resource Center (http://www.brc.riken.go.jp/Eng/index.html).
我们利用局部转座系统，将拟南芥（诺森生态型）第5号染色体上三个起始位点的解离（Ds）元件进行了转座。我们确定了Ds元件侧翼的部分基因组序列，并通过与已发表的基因组序列进行比较，在1173个转座系中定位了这些元件的插入位点。大多数品系含有单个Ds元件拷贝。其中一半品系的Ds元件位于第5号染色体上；特别是，在三个起始位点附近明显观察到插入“热点”。在其他品系中，Ds元件跨染色体进行了转座。我们分别在第2号和第4号染色体的顶端，靠近核仁组织区2和4的位置发现了其他插入热点。另一个特征是，Ds元件倾向于在染色体末端附近转座，而很少在着丝粒附近转座。尽管这三个起始位点拥有相同的Ds构建体，但其分布模式仍存在差异。超过一半的Ds元件不规则地插入到基因组中；也就是说，它们既没有保留Ds的完美反向重复序列，也没有留下完美的靶位点重复序列。这种对分布模式的精确分析将有助于全面了解转座机制。基于这些Ds插入位点，我们构建了一个数据库，用于在计算机上筛选基因敲除突变体。在1173个品系中的583个品系中，Ds元件插入到了蛋白质编码基因中，这表明这些品系是基因敲除突变体。该数据库和各个品系将可从日本理化学研究所生物资源中心（http://www.brc.riken.go.jp/Eng/index.html）免费获取，供学术使用。
