Expression analysis of gene trap lines and mapping of donor loci for Dissociation transposition in Arabidopsis

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In order to establish a system for characterizing gene function utilizing information obtained from genome sequences, we generated T-DNA insertion lines using a newly constructed binary vector. The vector carries a uidA [β-glucuronidase (GUS)] reporter gene which allows the promoter activity of the inserted genes to be monitored, a transposable element Dissociation (Ds) for targeted insertional mutagenesis, and the cis sequences required for Agrobacterium-mediated transformation. Approximately 8% of the 20,000 lines tested for GUS activity exhibited positive staining. Staining was detected in various organs including roots, leaves, stems, flowers, and siliques. These lines are therefore useful resources for analyzing tissue or organ specific gene expression. We have included the GUS expression patterns on our web site (http://www.kazusa.or.jp/ja/plant/GUS/). To establish a system for targeted insertional mutagenesis, integration sites of the T-DNAs in 140 lines with a single T-DNA insertion were mapped on the genome. The T-DNA contains the Ds element; therefore these 140 lines could be used as donor loci for Ds transposition. The integration sites were almost evenly distributed on all five chromosomes except for the nuclear organizer regions of chromosomes 2 and 4 and the centromeric regions. The Ds element in one line was transposed in combination with an Activator (Ac) element. In about half of the transposed lines, the Ds elements were reinserted within about 1 M bp from the donor locus. These results indicate that the donor loci for Ds transposition that were mapped in this study are a valuable resource for targeted mutagenesis throughout the Arabidopsis genome.

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