G02004: Development and comparison of molecular profiling methods for improved safety evaluation using GM Brassicas
Wednesday 16 October 2002
This research project aims to advance molecular profiling technologies with the aim of testing their feasibility for practical application in the safety evaluation of GM foods.
Background
This project aims to advance molecular profiling technologies with the aim of testing their feasibility for practical application in the safety evaluation of GM foods. The robustness of 'substantial equivalence' in safety evaluations could be advanced by these technologies but they have not been fully explored or validated for this purpose.
Research Approach
The project includes the three major profiling strategies: metabolite profiling; proteomics; DNA microarrays. It intends to explore their relative merits in detail using common study materials. The project is focussed on Brassicas, chosen to provide the combined advantages of crop plants likely to be prominent in the application of second generation GM technology and the advanced genomic characterisation of Arabidopsis. The GM traits chosen for study represent a range of existing and future applications that are typical of agronomic, quality and functional targets. An important component of this study is the inclusion of an analytical framework to relate differences in a GMO to the background of natural genetic variation. This is essential for the practical application of these technologies in safety evaluations.
Results and findings
This project investigated the application of transcriptomics, proteomics and metabolomics techniques, such as DNA microarrays and 2D gel electrophoresis, to analysing brassicas, using arabidopsis as a model plant system.
Project G02004 differed from the other G02 projects in that a principle objective was to use the 'omics techniques to analyse a number of independent lines carrying the same transgenes but with vastly differing levels of expression. This work was undertaken in the model plant Arabidopsis thaliana because its genome has been sequenced thereby providing detailed molecular data. A second objective was to determine the transferability of the methods to another brassica species (broccoli) that is grown as a food crop.
It was possible to demonstrate that data produced by each of transcriptomics, proteomics and metabolomics techniques could be integrated providing a complete biological picture.
During the course of the project a new approach, Multi-dimentional Protein Identification Technology (MudPIT), became available. MudPIT was demonstrated to be highly sensitive with more than 300 proteins, from crude extracts of conventional Arabidposis leaves, being identified by amino acid sequence.
No unintended proteomic effects were seen in transgenic arabidopsis plants expressing the 'neutral' gene GUS. In plants expressing a gene for a novel enzyme (HCHL) that is metabolically active, the extent of the changes in RNA and protein species and metabolites is directly proportional to the level of expression of the transgene. The activity of the HCHL led to the synthesis of new metabolites in arabidopsis. It was also demonstrated that changing light levels lead to very significant differences in the numbers of genes expressed.
In all the crops analysed, environmental stress had a much greater impact on the transcriptome, proteome and metablome than transgene introduction.
Dissemination information
The final report is available from the FSA Information Centre.
To obtain a copy, please contact the Enquiry Desk, Information Services, Food Standards Agency (tel: 020 7276 8181/8182 or email: library&info@foodstandards.gsi.gov.uk)