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Unintended Effects of Genetic Manipulation
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Discoloration and DNA rearrangements were observed in transgenic tobacco plants expressing HIV proteins.

Manipulated Organism: Tobacco (Nicotiana tabacum).

Inserted Transgenes: Genes for the HIV proteins p24 and Nef, as well as for the fusion proteins p24-Nef and Nef-p24, were inserted in the chloroplast genome under the control of the ribosomal RNA promoter (Prrn). The antibiotic resistance gene aadA was also included as a selectable marker.

Goal of This Study: Tobacco plants are being engineered as potential "bioreactors" for producing pharmaceuticals. The goal of this study was to create transgenic tobacco plants that accumulate high levels of the HIV proteins p24 and Nef, for possible use in an HIV vaccine.

Intended Effect: Tobacco lines were created that accumulate p24 and p24-Nef in young leaves, at concentrations as high as 40% of the total soluble protein.

Results of This Study:
  • Transgenic expression of p24 and p24-Nef was variable across leaves of different ages. There was a trend toward less transgenic protein in older leaves.

  • Most plants with the Nef transgene had normal color and vigor, but these turned out to have negligible amounts of Nef protein because of unexpected DNA rearrangements that silenced Nef expression. The few transgenic plants that did express Nef were yellow and slow-growing.

  • The transgenic fusion protein Nef-p24 was detectable but appeared to have been partially degraded.

  • The fact that attempts to express Nef and Nef-p24 at high levels were foiled while p24-Nef worked well suggested that the front end (N-terminus) of Nef interfered with normal chloroplast function.

Source: Zhou, F., J. A. Badillo-Corona, D. Karcher, N. Gonzalez-Rabade et al. (2008). "High-level Expression of Human Immunodeficiency Virus Antigens from the Tobacco and Tomato Plastid Genomes," Plant Biotechnology Journal vol. 6, pp. 897-913.

Author Affiliations: Author Affiliations: Max Planck Institute for Molecular Plant Physiology, Potsdam-Golm, Germany; University of Cambridge, UK; Trinity College, Ireland.

Funding: European Union FP6 Grant; Max Planck Society.

Product Status: Not on the market as of 2009.

Copyright 2009 The Nature Institute.

This document: http://natureinstitute.org/nontarget/reports/tobacco_002.php

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