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  • 1
    ISSN: 1573-5028
    Keywords: plants ; antisense ; suppression ; nopaline synthase ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We report the successful suppression of nopaline synthase (EC 1.5.1.19) enzymatic activity in the leaves of tobacco plants via the overproduction of RNAs complementary to the nopaline synthase (nos) mRNA. Several different regions of the nos gene were fused, in antisense orientation, to the promoter from a strongly expressed petunia chlorophyll a/b-binding protein gene. These constructions were directly introduced into a tobacco line which contained a single copy of the wild-type nos gene and transgenic plants were regenerated. The degree of nopaline synthase suppression in the leaves of the double transformants ranged up to 85% and was dependent on the particular region of the nos gene present in the antisense RNA. The most effective nos antisense sequences were derived from the 3′ half of the nos gene transcript. In addition, we report a new sensitive method for the detection and quantitation of nopaline synthase activity in crude plant extracts.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1617-4623
    Keywords: Promoter strength ; Transformed callus ; Chitinase gene ; Octopine synthase gene ; Agrobacterium
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The 35S promoter of cauliflower mosaic virus and promoters from the nopaline synthase, 1′ and 2′ genes of Agrobacterium tumefaciens T-DNA were fused to the bacterial octopine synthase and chitinase gene coding regions. These chimaeric gene constructions were introduced into tobacco, sugarbeet and oilseed rape cells and their relative levels of expression measured by primer extension analysis of RNA isolated from pooled populations of stably transformed calli. In tobacco callus, the 35S promoter provided the highest levels of gene expression, followed by the 2′, 1′ and nopaline synthase promoters. While the ranking of these promoters is conserved in sugarbeet and oilseed rape callus, there is between-species variation in the relative strength of these promoters. In all three species, transcription initiation is conserved for each of the chimaeric gene constructions. Additional constructions in which the 5′ untranslated leader of a petunia chlorophyll a/b binding protein gene is substituted for DNA downstream of the 35S transcription start site demonstrates that heterologous 5′ leader sequences can be utilized to augment steady-state levels of reporter gene expression
    Type of Medium: Electronic Resource
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