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        Promoter에 따른 밀 소포자 유래 배에서 transient GUS 유전자 발현

        김경문,송인자,P.Stephen Baenziger 한국육종학회 2007 한국육종학회지 Vol.39 No.3

        Transient expression profiles for several chimeric β-glucuronidase (GUS) gene constructs were determined in microspore-derived embryos of wheat following microprojectile bombardment. The constructs analyzed consisted of the uidA (GUS) reporter gene driven by six different promoters [Cauliflower Mosaic Virus 35S (CaMV35S), Nopaline synthase (NOS), Mannopine synthase (MAS), Chlorella Mosaic Virus Adenin methyltransferase (AMT), maize Ubiquitin 1 (UBI1), and enhanced 35S (E35S)]. The total numbers of GUS blue spot were determined manually under a dissecting microscope after histochemical staining for GUS. Results suggest that the E35S promoter is the most active and UBI1 promoter is the second active in embryos or embryogenic calli derived from wheat microspore. In addition, by flurometric assay on GUS, the E35S promoter was the best. Therefore, both UBI1 and E35S promoter are suitable for constitutive expression of the gene of interest in microspore-derived embryos of wheat. This information describing promoter functionality in wheat will be important when designing gene constructs for traits modification and when choosing appropriate cultivars for improvement through gene transfer experiments.

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        Effect of Osmoticum on GUS Transient Expression in the Bombarded Microspore-derived Embryos of Wheat(Triticum aestivum L.)

        Kyung Moon Kim,P. Stephen Baenziger,Thummala Chandrasekhar,Hyo Yeon Lee 한국육종학회 2007 한국육종학회지 Vol.39 No.2

        The effect of osmotic condition on β-glucuronidase (GUS) transient expression was evaluated in microspore-derived embryos of wheat. Microspore explants were treated on medium containing various mannitol concentrations prior to and post bombardment with plasmid DNA pAHC25 containing uidA gene controlled by maize ubiquitin 1 (UBI1) promoter. GUS expression in the bombarded explants was examined by histochemical and fluorometric assays. Increased GUS expression was observed with mannitol treatment when compared to untreated explants. The histochemical study showed that the number of blue (GUS) foci were the highest in the bombarded explants treated with 0.6 M mannitol medium. The fluorometric assay of bombarded explants also proved 3.5-fold increase in GUS activity with 0.6 M mannitol treatment when compared to without mannitol treatment. These results indicate that 0.6 M mannitol is beneficial for improving transformation efficiency of wheat microspore-derived embryos or embryogenic calli through biolistic transformation.

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