Tobacco was transformed with the nitroreductase (<i>nfsI</i>) gene from<i> Enterobacter cloacae. </i>Nitroreductase (NR) was proposed to reduce TNT to classic type I nitroreductase products and its activity was found to be induced in the presence of TNT. The aim of this study was to characterize these plants with respect to their ability to tolerate, take up and transform TNT from their environment. The product/s of TNT reduction by nitroreductase has not been characterized prior to this study but were analyzed here and found to be 4-hydroxylamino-2,6-dinitrotoluene (4-HADNT). Semi-quantitative RT-PCR analysis of transgenic lines indicated that higher <i>nfsI </i>transcript expression was correlated with greater tolerance to TNT at germination. The most TNT-tolerant transgenic line was established and chosen for further characterization in sterile, aqueous conditions and in soil contaminated with TNT. Vegetative growth was suppressed in wild type plants at 23mg/L TNT whereas transgenic plants were found to remove all TNT from a concentration of 113mg/L TNT and were found to gain biomass at this concentration. This concentration of TNT is above the aqueous solubility limit of TNT and would be presumably be the highest concentration plants would encounter in the field. Following uptake, transgenic seedlings transformed TNT predominantly to 4-HADNT. This product was only detected in the tissue and growth medium of transgenic plants and its high levels appeared to correlate with enhanced tolerance and transformation of TNT. The wild type seedlings produced both isomers of the further reduced product of TNT, the amino-dinitrotoluene (2-ADNT and 4-ADNT). This may be significant, as the 2 isomer of TNT reduced products have been found to be more toxic to other organisms. The phytodetoxification of TNT was also studied with both plant lines in TNT-contaminated soil. The transgenic seedlings were found to tolerate TNT significantly better than the wild type plants at germination and during vegetative growth.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:603658 |
| Date | January 2003 |
| Creators | Hannink, N. K. |
| Publisher | University of Cambridge |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
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