Establishment of an enhancer trap system with Ds and GUS for functional genomics in rice.
To develop an efficient means of enhancer trapping, a two-element system employing Ds and an Ac transposase (AcTPase) gene was tested in rice. We generated 263 transgenic rice plants, each of which harboured the maize transposable element Ds together with a GUS coding sequence under the control of a minimal promoter (Ds-GUS), and a gene that confers resistance to the herbicide chlorsulfuron. Among the 263 lines generated, 42 were shown to have a single copy of the Ds-GUS element. Four single-copy lines were crossed with each of six transgenic plants that carried the AcTPase gene. Excision of the Ds-GUS in leaves of F1 plants was detected in eight combinations out of seventeen examined. The frequency of transposition of Ds-GUS in germ cells in the F1 plants was examined using 10,524 F2 plants, and 675 (6%) were judged to be transposants. Their frequencies differed among F1 plants depending on the AcTPase x Ds-GUS cross considered, and also among panicles on the same F1 plant. This suggests that Ds-GUS tends to transpose during panicle development. Southern analysis with a GUS probe showed different band patterns among transposants derived from different panicles. Therefore, the transposants derived from different panicles must have arisen independently. Transposants showing tissue-specific GUS activities were obtained, and enhancers thus trapped by the Ds-GUS element were identified. These results demonstrate that the system is suitable for the isolation of large numbers of independent Ds-GUS transposants, and for the identification of various tissue-specific enhancers. The Ds-GUS lines generated in this study offer a potentially powerful tool for studies on the functional genomics of rice.
Ito
Y
Y
Plant Genetics Laboratory, National Institute of Genetics, 1111 Yata, Mishima, Shizuoka-ken 411-8540, Japan.
Eiguchi
M
M
Kurata
N
N
D015139:Blotting, Southern; D003433:Crosses, Genetic; D004251:DNA Transposable Elements; D004351:Drug Resistance; D004742:Enhancer Elements, Genetic / Q000502:physiology*; D018506:Gene Expression Regulation, Plant; D005787:Gene Frequency; D018745:Genome, Plant; D005966:Glucuronidase / Q000235:genetics / Q000378:metabolism; D006540:Herbicides / Q000633:toxicity; D012275:Oryza / Q000235:genetics* / Q000254:growth & development; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic; D013449:Sulfonamides / Q000633:toxicity; D019895:Transposases; D014227:Triazines / Q000633:toxicity; D003313:Zea mays / Q000235:genetics* / Q000254:growth & development
D004251:DNA Transposable Elements; D006540:Herbicides; D013449:Sulfonamides; D014227:Triazines; D019895:Transposases; D005966:Glucuronidase; C037137:chlorsulfuron