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Tobacco ribosomal DNA spacer element stimulates amplification and expression of heterologous genes

Nature Biotechnology volume 18, pages 1303–1306 (2000)Cite this article

Abstract

Here we show that the cis-acting genetic element aps (amplification-promoting sequence), isolated from the nontranscribed spacer region of tobacco ribosomal DNA (rDNA), increases the level of expression of recombinant proteins. Transgenic tobacco plants, transformed with expression cassettes containing the herbicide-resistant acetolactate synthase (hr-ALS) gene or the green fluorescent protein (GFP) gene fused to the aps sequence, had greater levels of corresponding messenger RNAs (mRNAs) and proteins compared to transformants lacking aps. Analysis of transgenic plants showed that aps increased the copy number and transcription of the adjacent heterologous genes and, in the case of hr-ALS, enhanced the herbicide resistance phenotype. Both the increased transgene copy number and enhanced expression were stably inherited. These data provide the first evidence that the aps sequence can be used for gene amplification in transgenic plants and possibly other multicellular organisms.

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Figure 1: Identification and molecular organization of the tobacco aps.
Figure 2: Expression of the hr-ALS gene in ALS and ALS-aps transformants.
Figure 3: Expression of GFP in GFP and GFP-aps transformants.

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Acknowledgements

We thank C. Reichel for the GFP expression cassette and F. Petersen for help with GFP detection. hr-ALS expression cassette, and the samples of Pursuit herbicide were kindly provided by the American Cyanamid Company (Princeton, NJ). This work was supported by a grant from Phytomedics, Inc. (Dayton, NJ).

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Author notes

  1. Ludmyla Borisjuk

    Present address: Institute of Plant Genetics, Correns Str. 3, Gatersleben, D-06466, Germany

Authors and Affiliations

  1. Biotech Center, Foran Hall, Cook College, Rutgers University, 59 Dudley Rd., New Brunswick, 08901-8520, NJ, USA

    Nikolai Borisjuk, Slavko Komarnytsky, Sofia Timeva & Ilya Raskin

  2. Phytomedics Inc., 2245 Route 130, Suite 103, Dayton, 08810, NJ, USA

    Nikolai Borisjuk & Ludmyla Borisjuk

  3. Center of Plant Molecular Biology, University of Tuebingen, Auf der Morgenstelle 28, Tuebingen, D-72076, Germany

    Vera Hemleben

  4. Institute of Cell Biology and Genetic Engineering, Zabolotnogo St., 148, Kyiv, DSP-22, 252650, Ukraine

    Yuri Gleba

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Correspondence to Ilya Raskin.

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Borisjuk, N., Borisjuk, L., Komarnytsky, S. et al. Tobacco ribosomal DNA spacer element stimulates amplification and expression of heterologous genes. Nat Biotechnol 18, 1303–1306 (2000). https://doi.org/10.1038/82430

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