The Best Kept Secret of GM Crops

For ACRE open hearing on the criticisms of T25 GM maize risk assessment

The hearing will take place from 10.00am to 2.00pm on Wednesday, 20
February, in Room 7A, B and C, Ashdown House, Department for Environment, Food and Rural Affairs, 123 Victoria Street, London SW1E 6DE.

Dr. Mae-Wan Ho

Institute of Science in Society, PO Box 32097, London NW1 0XR, UK

I am speaking against the market approval of T25 because there is no evidence that it is a genetically stable, uniform line, the single most important criterion for approval. For unless it is genetically stable, you might as well forget about environmental or health risk assessment. And genetic instability is also a serious safety issue. The public hearing on T25 was suspended over a year ago when it was found not to have passed the required EC test for Distinctiveness, Uniformity and Stability (the DUS test), as I pointed out when giving evidence to the hearing [1].

The new EC Directive on deliberate release requires strict molecular evidence of genetic stability, which is also necessary for establishing the identity of the transgenic line and to ensure traceability. The best-kept secret of GM crops is that they are not stable.

There is a large literature on gene silencing, in which the transgenes remain in the genome, but are not expressed. More serious, from the safety point of view, is structural instability, the tendency for the transgenic DNA to come loose, to rearrange or become lost in part or in whole in successive generations [2,3]. This could change the transgenic line in unpredictable ways in terms of health and environmental risks. And it will increase the chance of transgenic DNA being taken up by unrelated species to make new combinations with their genetic material. That’s referred to as horizontal gene transfer and recombination. Transgenic DNA can spread to every species that interact with the transgenic plant, in the soil, in the air, in the mouth and gut and the respiratory tracts of animals including human beings.

New viruses and bacteria that cause diseases could be generated, and antibiotic resistance marker genes could spread to the pathogens. Transgenic DNA may also get into human cells and insert into the human genome; and a large body of evidence from so-called gene therapy experiments have amply demonstrated this does occur [4]. The constructs used in gene therapy are very similar to those used in transgenic plants, and one main side-effect of transgenic DNA inserting into human genome during gene therapy is cancer.

Despite that, our regulators have not required biotech companies to provide molecular evidence of stability. ACRE’s latest guidelines for industry put out for public consultation asks industry to provide molecular evidence of genetic stability over one generation only [5], which is derisory. We need data for at least five successive generations [6]. No such data have come forward from the companies. On the contrary, companies have been allowed to hide under ‘commercial confidentiality’.

I am putting to you twelve reasons why trangenic DNA is different from natural DNA, and is more likely to spread by horizontal gene transfer and recombination, both by design and otherwise. I hope you will refute these point by point.

(The details are in two ISIS reprint collections on transgenic instability and horizontal gene transfer that I am presenting to ACRE, for free.)

• All artificial constructs tend to be unstable, so much so that this is a topic in a standard text-book on genetic engineering [7]. Transgenic DNA is more likely to break and join up again, ie, to recombine.
• Transgenic DNA typically contains DNA from widely different sources, mainly bacteria and viruses and other genetic parasites that cause diseases and spread antibiotic resistance, and hence, has the potential to recombine homologously with all those agents, ie, due to similarities in DNA base-sequence. Homology enhances horizontal gene transfer 10 million to 100 million-fold [8].
• Transgenic DNA is designed to cross species barriers and to invade genomes. They are flanked by recombination sequences, such as the left and right borders of T-DNA or the terminal repeats of viral vectors, which enable them to jump into genomes. By the same token, they could jump out again. Enzymes catalysing jumping in also catalyse jumping out.
• Certain ‘receptive hotspots’ have now been identified in both the plant [9] and the human genome [10]. These may also be ‘recombination hotspots’, prone to breaking and rejoining. That would mean inserted transgenes are more likely to be lost, to recombine, or to invade other genomes.
• There are mechanisms in the cell that actively seek out, inactivate or eliminate foreign DNA from the genome [11].
• Cell and embryo culture methods are well-known to induce unpredictable, uncontrollable (somaclonal) variations that persist in the plants generated. There is now evidence that the transformation process for making transgenic plants induces further genetic instability [12-14] leading to chromosomal rearrangements, genome scrambling, in other words.

• Monsanto’s Roundup Ready soya, commercially grown for years, was finally analysed by molecular methods. Not only is the gene order of the insert found to be scrambled, the plant genome at the site of insertion is also scrambled, and there is a 534 bp fragment of unknown origin in there as well [15]. All very different from the original data provided by Monsanto.

• Recombination hotspots within the transgenic DNA, such as that associated with the ubiquitous cauliflower mosaic virus (CaMV) 35S promoter, could enhance horizontal gene transfer and recombination. We highlighted that in 1999 [16-18], and demanded that all transgenic crops with the promoter should be immediately withdrawn for safety reasons. Two years later, the researchers who discovered the promoter’s recombination hotspot also recommended that it should no longer be used [19], not because of safety, but because its instability compromises agronomic performance.

• Recently, landraces of corn growing in remote regions of Mexico were found contaminated with transgenic corn DNA by probing with the CaMV 35S promoter [20]. Molecular analysis showed that the sequences next to the promoter are very diverse, as consistent with horizontal gene transfer and recombination [21].
• CaMV 35S promoter is active in species across the entire living world, including frog eggs and human cells [18], as we uncovered in the literature more than ten years old that had apparently escaped the notice of plant geneticists who attacked us. CaMV 35S promoter, if transferred to human or animal cells, could activate cancer-associated genes as well as dormant viruses that are in all genomes. Another side effect of gene-therapy is the generation of active viruses in cell lines used to package the gene-therapy vectors [4]. Our critics are still dismissing the risks of CaMV 35S promoter, but are avoiding doing any experiments. It is a case of don’t look, don’t see [5].
• Transgenic DNA from GM plants was found to transfer to soil bacteria. The possibility of transfer to bacteria in the mouth and gut of animals was suggested in laboratory investigations funded by the UK government. There is also evidence suggesting that transgenic DNA from crop plants has transferred to soil bacteria in the field [22]. But ACRE has ignored that by a selective interpretation of the scientific evidence that seems to me contrary to both the precautionary principle and good science [23].

In summary, there is no reason to believe T25 is stable. Furthermore, it has especially hazardous sequences, including the CaMV 35S promoter and an ampicillin resistance gene that, though inactive, can easily be transferred into integrons that will provide it with a promoter to make it functional [1]. T25 has uncharacterised sequences that might be involved in causing diseases. Finally, it has an origin of replication, which enables the transgenic DNA to be replicated as a plasmid if transferred into bacteria, thereby greatly increasing horizontal gene transfer on to other species. The origin of replication is also a recombination hotspot, and there have been strong recommendations from a recent joint FAO/WHO Expert Consultation on Foods Derived from Biotechnology that transgenic lines containing this sequence should not be approved on safety grounds [24].

1. Ho MW. Chardon LL Public Hearing Ocober 26 2000 on behalf of Burnham Group, also in transcript.
2. See Ho MW. Genetic Engineering Dream or Nightmare? Gateway, Gill & Macmillan, Bath and Dublin, 1998, 1999, Chapter on Perils amid Promises of Genetically Engineered Foods.
3. ISIS Reprints on Transgenic Instability, 1999-2001, ISIS Publications, London.
4. Ho MW, Ryan A, Cummins J and Traavik T. Slipping Through the Regulatory Net: ‘Naked’ and ‘Free’ Nucleic Acids, Third World Network Biotechnology Series, Third World Network, Penang 2001.
5. See Watering down EC Directive on Deliberate Release ISIS Report, February 2002.
6. Ho MW and Steinbrecher RA. Fatal flaws in food safety assessment: critique of the joint FAO/WHO Biotechnology and Food Safety Report. Environmental & Nutritional Interactions 1998, 2, 51-84.
7. ISIS Reprints on Horizontal Gene Transfer, 1999-2001, ISIS Publications, London.
8. Principles of gene manipulation, by Old and Primrose, Blackwell Science, 5^th ed, 1994.
9. DeVries J, Meier P and Wackernagel W. The natural transformation of the soil bacteria Pseudomonas stutzeri and Acinetobacter sp. by transgenic plant DNA strictly depends on homologous sequences in the recipient cells. FEMS Microbiology Letters 2001, 195, 211-5.
10. Kumar S and Fladung M. 2000. Transgene repeats in aspen: molecular characterisation suggests simultaneous integration of independent T-DNAs into receptive hotspots in the host genome. Mol Gen. Gent 2000, 264, 20-8.
11. Miller DG, Rutledge EA and Russell DW. Chromosomal effects of adeno-associated virus vector integration. Nature genetics 2002, 30, 147-8.
12. Kumpatla, S.P., Chandrasekharan, M.B., Iyer, L.M., Li, G. and Hall, T.C. (1998). Genome intruder scanning and modulation systems and transgene silencing. Trends in Plant Sciences 3, 96-104.
13. Horvath H, Jensen L,Wong O, Kohl E, Ullrich S, Cochran J, Kannangara C, and von Wettstein D. Stability of transgene expression, field performance and recombination breeding of transformed barley lines, Theor Appl Genet. 2001,1-11.
14. Svitashev S, Ananiev E, Pawlowski WP, and Somers DA. 2000. Association of transgene integration sites with chromosome rearrangements in hexaploid oat. Theoretical and Applied Genetics 2000, 100,: 872-80.
15. Tax FE and Vernon DM. T-DNA-associated duplication/transloations in Arabidopsis. Implications for mutant nanalysis and functional genomics. Plant Physiology 2001, 126, 1527-38.
16. Windels P, Taverniers I, Depicker A, Van Bockstaele E and De Loose M (2001). Characterisation of the Roundup Ready soybean insert. Eur Food Res Technol DOI 10.1007/ s002170100336, © Springer-Verlag; see also "Scrambled genome of Roundup Ready soya" by Mae-Wan Ho, ISIS Reprints on Transgenic Instability, 1999-2001, ISIS Publications, London.
17. Ho MW, Ryan A and Cummins J. Cauliflower mosaic viral promoter – a recipe for Disaster? Microbial Ecology in Health and Disease 1999: 11: 194-197.
18. Ho MW, Ryan A and Cummins J. Hazards of transgenic plants with the cauliflower mosaic viral promoter. Microbial Ecology in Health and Disease 2000: 12: 6-11.
19. Ho MW, Ryan A and Cummins J. CaMV35S promoter fragmentation hotspot confirmed and it is active in animals. Microbial Ecology in Health and Disease 2000: 12: 189.
20. Christou P, Kohli A, Stoger E, Twyman RM, Agrawal P, Gu X. Xiong J, Wegel E, Keen D, Tuck H, Wright M, Abranches R and Shaw P. Transgenic plants: a tool for fundamental genomics research. John Innes Centre & Sainsbury Laboratory Annual Report 1999/2000, p. 29. See "Top research centre admits GM failure" ISIS Reprints on Transgenic Instability, 1999-2001, ISIS Publications, London.
21. Quist D and Chapela IH. Transgenic DNA introgressed into traditional maize landraces in Oaxaca, Mexico. Nature 2001, 414, 541-3, 2001.
22. "Transgenic pollution by horizontal gene transfer?" by Mae-Wan Ho, in ISIS Reprints on Horizontal Gene Transfer, 1999-2001, ISIS Publications, London.
23. "Horizontal gene transfer happens. A practical exercise in applying the precautionary principle" by Mae Wan Ho in ISIS Reprints on Horizontal Gene Transfer, 1999-2001, ISIS Publications, London.
24. Joint FAO/WHO Expert Consultation on Foods Derived from Biotechnology, WHO Headquarter, Geneva, September 24-28, 2001.