Is it or isn't it?

Cisgenesis, zinc finger nuclease technology, reverse breeding, oligonucleotide-directed mutagenesis etc – the field of new techniques is broad and complex. New techniques or adaptations of existing techniques are constantly being added. The current political discussion, and the debate within public authorities or critics of genetic engineering, mostly focus on whether a particular technique or the product derived from it should be classed as genetic engineering or not, using the definition of genetic engineering set out in Art. 2.2 of the Release Directive (2001/18/EC) as a basis. In order to avoid too much technical detail, it is helpful to offer an initial categorisation of techniques based on their respective approach.

Category 1: Despite all claims to the contrary: "Classic" genetic engineering

Many of the new techniques are not really new, but correspond to techniques that have been established for over 20 years. This applies to both the breeding process and end products. Presently, attempts are being made to describe these techniques and their resulting products as conventional forms of breeding. Sometimes this even employs arguments brought to bear by the critics of genetic engineering. It is argued, for example, that unlike transgenesis, cisgenesis does not cross the species barrier and that the same results could therefore also be achieved through conventional breeding. The methods of transformation, however, clearly do represent forms of genetic engineering (particle bombardment or Agrobacterium tumefaciens). Even if the new gene originates from a species compatible for cross-breeding, it is impossible to predict where it will be integrated in the genome. This is what constitutes the risk inherent in this technique, in contrast to conventional breeding.

This category particularly includes the following procedures: Cisgenesis, intragenesis, floral dip and the use of genetically modified scions. Grafting onto genetically modified rootstock for commercial growing (not just during the breeding process) also falls within this category. It is incorrect to claim that harvested products (such as apples) derived from a scion grafted onto a GM parent plant do not constitute GMO. It is possible, for example, that proteins from the GM rootstock are transported to the non-GM scion; the phenotype of the scion and its product could therefore be altered. We believe the position of the Central Commission for Biological Safety (Zentrale Kommission für die Biologische Sicherheit, ZKBS), as outlined in their statement, to be unjustified; this demands that only GM rootstock should be classified as GMO and not the resulting harvested products (ZKBS, 2012:10). Even if no traces of transgenic or cisgenic DNA are found in the product, the principle of process-based evaluation which currently prevails in Europe mandates that the entire organism should be regulated as GMO, both for the purpose of growing these organisms and for labelling the resulting harvested crops.

Category 2 – "Classic" genetic engineering processes that do not result in GMO?

During the breeding process, techniques of transgenesis and cisgenesis are used to directly alter the plant. This takes place, for example, during reverse breeding, agro-infiltration (except for floral dip), or fast track breeding. Either a GM host is used during the breeding process to induce early flowering, or GE methods are combined with the crossbreeding of different parent plants. The foreign genes transferred to induce early flowering can originate from other varieties of the same species, local cultivars, or related wild varieties, but also from species that are not normally crossable (such as birch in the case of fast-track apple breeding). The intention is to speed up the breeding process by actively introducing the new trait of early flowering, achieved by crossing an early flowering GM host with another, non-GM variety followed by backcrossing. The end product – the new variety – no longer contains any DNA of the GM parent plant. Due to the principle of process-based evaluation prevailing in Europe, and many open questions with respect to risk evaluation, we are of the opinion – which is shared by others – that the techniques in this category should not just be evaluated based on their respective process (which unquestionably uses GM plants), but that the resulting breeding products should also be regulated as GMO, even if they no longer contain traceable evidence of transgenesis or cisgenesis.

Category 3: New techniques which directly interfere with DNA at the molecular level and/or gene regulation

This category primarily comprises the following techniques: Zinc finger nucleases (ZFN 1-3), TALEN techniques, meganucleases, Oligonucleotide Directed Mutagenesis (ODM), CRISPR-Cas, RNA-directed methylation (RdDM) and RNAi technology.

We present RNAi as an example. This technique, which is used among others by Monsanto, heavily relies on double-stranded RNAs. Put very simply, these double-stranded RNAs can selectively switch off genes by inhibiting an intermediate stage of gene expression. Companies are using this process by genetically modifying plants to synthesise insect-specific RNAs. When certain pests feed on the plant, they ingest these RNAs, which then turn off specific genes essential to the survival of the insect and kill it. So far, RNAi has not been used successfully in human therapy as it has proven difficult to transfer RNAs to human cells. In contrast, insects and especially their voracious larvae easily absorb ingested RNAs in their midgut, from which they can spread throughout the body. The new insecticide RNAs are supposed to be specific enough to not affect even closely related insect species.

Presently, plant seed is being tested in the USA which has been modified to enable resistance to western corn rootworm. The RNA used turns off the gene responsible for Snf7. Snf7 helps to transport proteins to their proper place in cells; without this function beetle larvae die within a few days. But how can we be sure that this anti-Snf7-RNA is not also dangerous to other animals or even humans? Critics see a problem in this, referring to the work of Zhang et al. (2012) which demonstrates that small RNAs from food plants can be traced in the blood of mice and humans. The consequences of this have remained unclear so far, not least because very little (independent) risk research is being carried out in this field.

The techniques of category 3, which have also been termed "synthetic genetic engineering" (Testbiotech 2013), are particularly suited to illustrate the complexity of the topic and the rapid change that is taking place within science. Both aspects render the topic of "new molecular techniques" complicated and largely inaccessible to public debate – a debate which is nevertheless urgently needed. On the other hand, these techniques demonstrate a certain continuity in the development of plant breeding from conventional breeding through to genetic engineering and finally the new molecular techniques: From populations to varieties, from varieties to traits, from traits to constituents, from constituents to genes, from genes to single nucleic acids, and now from single nucleic acids to the methyl groups of single nucleic acids – the depth of intervention is growing constantly. This is linked to far-reaching contextual shifts, both in the practice of breeding, which is increasingly becoming laboratory-based, and in the corresponding cultivation systems.

References:

Testbiotech 2013: Stellungnahme Synthetic Genome Technologies. Download: Testbiotech

Verbändepapier (AbL, BUND, GeN, Greenpeace, IG Saatgut, Save our Seeds, Testbiotech, Zukunftstiftung Landwirtschaft) 2014: Neue gentechnische Verfahren in Pflanzen- und Tierzucht müssen reguliert werden. Stellungnahme aus Anlass des Symposiums „Herausforderungen 2015: Neue Entwicklungen in der Gentechnik – Neue Ansätze für das behördliche Handeln?“ des Bundesamtes für Verbraucherschutz und Lebensmittelsicherheit (BVL), 5.-6. November 2014, Berlin.

Zentrale Kommission für die Biologische Sicherheit (ZKBS) 2012: Stellungnahme der ZKBS zu neuen Techniken für die Pflanzenzüchtung.

Zhang, L. et al. 2012: Exogenous plant MIR168a specifically targets mammalian LDLRAP1: evidence of cross-kingdom regulation by microRNA. In: Cell Research 22, pp. 107-126.