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Unformatted text preview: JARQ 39 (4), 269 – 274 (2005) http://www.jircas.affrc.go.jp 269 Efficient Transgene Expression in Chrysanthemum, Chrysanthemum morifolium Ramat., with the Promoter of a Gene for Tobacco Elongation Factor 1 α Protein Ryutaro AIDA 1,3 * , Shingo NAGAYA 2 , Kazuya YOSHIDA 2 , Sanae KISHIMOTO 1 , Michio SHIBATA 1 and Akemi OHMIYA 1 1 Department of Genetics and Physiology, National Institute of Floricultural Science (Tsukuba, Ibaraki 305–8519, Japan) 2 Graduate School of Biological Sciences, Nara Institute of Science and Technology (Ikoma, Nara 630–0192, Japan) Abstract We investigated the usefulness of the promoter of a gene for tobacco elongation factor 1 α protein ( EF1 α ) for transgene expression in the chrysanthemum ( Chrysanthemum morifolium Ramat.). The EF1 α promoter was fused to the β-glucuronidase gene ( gus ) and introduced into the chrysanthemum. We obtained 238 putative transformants and GUS assay of the leaves of the in vitro plants revealed that 29.0% (69/238) of the putative plants were GUS-positive. The plants in the greenhouse that were 20 months after regeneration still showed a GUS activity in their leaves and petals. The tobacco EF1 α promoter expressed the transgene more efficiently than the 35S promoter of Cauliflower mosaic virus and could be used for transgene expression in chrysanthemum. Discipline: Biotechnology Additional key words: Agrobacterium , tissue culture, transformation, ornamental plants Introduction Chrysanthemum is one of the most popular and important ornamental plants in the world. There have been many reports of genetic transformation in chrysanthemum 7,16 . Many researchers have used the β- glucuronidase gene ( gus ) driven by the 35S promoter of cauliflower mosaic virus (CaMV), abbreviated as 35S/ gus , as a reporter gene for investigating transgene expres- sion in chrysanthemum. The GUS activity levels recorded in transgenic chrysanthemum with the 35S/ gus transgene have been low, ranging from 10 to 160 17 , 30 to 240 13 , and 30 to 250 15 pmol 4-MU mg –1 protein min –1 . The 35S or modified 35S promoter has been used to express practical transgenes for modifying characters such as resistance to diseases 9,15,17,19 , resistance to insects 14 , and flower color 5,6 . In some of these attempts, the mRNAs 5 or protein 17,19 of the transgene could not be detected, even when the transgenes were inserted in the genome. These results suggest that the 35S promoter does not function efficiently in chrysanthemum. Recently, several reports have been published on efficient promoters for transgene expression in chrysan- themum with the gus as a reporter gene. The potato Lhca3.St.1 (encoding the apoprotein 2 of the light-har- vesting complex of photosystem I) promoter expressed high GUS activity in leaf, stem, pedicel and ray floret (mean activity in the leaves among GUS positive plants was about 25,000 pmol 4-MU mg –1 protein min –1 ) 2 . The chrysanthemum UEP1 (encoding ubiquitin extension protein) promoter also expressed high GUS activity in...
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- Spring '09
- molecular biology, Chrysanthemum, GUS, Gus Activity