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Transgenic plant is a plant that contains foreign gene or genes, or indigenous genes that had been altered, which have been introduced artificially into its genome by using biotechnology techniques known as genetic engineering. The gene of interest (GOI) also known as transgene may come from the same plant species or variety or from other species, or may come from unrelated kind of organisms such as bacteria or animals. Once a transgenic plant is developed and become stable, the transgene can be inherited along with the rest of plant genome through normal pollination or propagated vegatatively. Transgenic plants are also often referred to as genetically engineered (GE) plants or genetically modified organism (GMO).
Generally, in a transgenic plant will also contain a selectable marker gene and a promoter which are driven the inserted genes for the functioning in plant host genome besides consisting of the interest gene. In my current research, the Eksotika papaya plant will be genetically modified to delay or prolong fruit ripening stage. This transgenic papaya plant will contain gene responsible for the synthesise of the enzyme ACC (1-amino-cyclopropane-1-carboxylic acid) Oxidase. The plant will also contain a bacterial gene (nptII) conferring resistance to the antibiotics kanamycin. This nptII gene was used as selectable marker for the selection of stably transformed cells from the majority of non-transformed cells, allowing the long term selection of transformed cells or tissues on a medium containing the selection agent during regeneration. Through this selection system, the transformed plantlets that carrying the desired fruit ripening genes can be selected prior to acclimatize and transplant to the field. In addition, a short regulatory sequence, 35S promoter of the cauliflower mosaic virus (CaMV), is inserted together with the interest and selectable marker genes.
The potential risks of delayed ripening transgenic Eksotika papaya to human and environment may include in the following risks:
The presence of antibiotic resistance gene, kanamycin
The toxicity or allergenicity of transgenic delayed ripening papaya to human and other organisms due to unforeseen or unintended effects.
The establishing of weediness: Could this transgenic papayas be harmful to the environment because of inherent weediness or increased potential to become a weed species, and
Horizontal gene transfer, meaning the transfer of the introduced gene (transgene) from transgenic delayed ripening papaya to the other organisms. Could the new genes introduced into the GM papayas will be transferred to other non-GM papaya or to other organisms, with adverse consequences.
1. Selectable Marker Gene
The potential risk of eating fruits from transgenic plant is due to the presence of antibiotic resistant gene in the fruit cells. Antibiotic resistance gene is frequently used at several stages in the developing genetically engineered plants. In the case of my study, the gene nptII, conferring resistance to the antibiotic kanamycin was used. Antibiotic-resistance plays an important role in the selection of transformed tissue. By using a suitable and optimal concentration of an antibiotic in the screening process, the technologist would be able to select and pickup transformed tissues, ie the tissues that contain the nptII gene in their cells. Those tissues without the nptII gene would die-off. This process of selection would reduce the number of escapees (regenerant plants without GOI). In my research, 150 mg/L kanamycin was used and placed in the culture media and the selection process was carried out for four months, consecutively. This antibiotic resistance gene will end up in the genetically engineered plants, whose function is no longer required after the selection process was completed.
So far, there is no evidence has been found that food produced or made from genetically engineered crop having nptII gene, which are commercially available now are not safe to be eaten compared to fruits produced using conventional method. For example Papaya varieties that genetically engineered to resist papaya ringspot virus (PRSV) have been accepted for food use in the US since 1997 and in Canada since 2003 (Gonsalves, et al., 2004). Human and animals consumed large amount of DNA in their daily diet. This DNA come from various kinds of cells, either from plants and animals constituting as food, and also may come from any contaminating microorganism or viruses that may present in the food. We already have been exposed to these varieties of DNA daily. Most of DNA that we consume or get through our alimentary canal are degraded by digestive enzymes and so for this antibiotic case.
There are worries on the potential risk of the development of antibiotic resistance microorganism exposed to the antibiotics. Horizontal gene transfer of DNA is known to occur in nature (Kidwell, 1993) but the likelihood of DNA transferred from plant material in the digestive system to microbe has not yet been experimentally determined. It is expected that for such horizontal transfer to be possible, it would have to come from consumption of fresh food only, since most post-harvest processing procedure would degrade the plant's DNA. Also many evidence showed that most DNA is rapidly degraded by the digestive system before it has a chance to integrate into the microfloral in our digestive system. Nevertheless, very low levels of uptake of gene sized DNA into cells of the gastrointestinal tract have been detected (Duggan et al., 2000; Doerfler, 2000; Einspanier et al.,2001).
There is also concern that the use of antibiotic resistance gene could reduce the effectiveness of antibiotics taken by a person who at the same time also consume the transgenic papaya carrying the resistance gene for that antibiotic. But studies shown that, the chances of this happening was probably very low due to rapid digestion of inactivating enzymes produced by the transgenic resistance gene. Also, this papaya only harbors a kanamycin resistant gene, an antibiotic that is not commonly used to treat infection in human.
2. Toxicity or Allergenicity To Humans And Other Organisms
Another potential risk is the inserted gene can cause an allergic reaction. However some people are already allergic to certain protein and they could have the allergic reaction if they are exposed to that protein from either conventional or transgenic products. In case of papaya delayed-ripening, the inserted gene comes from the plant itself and due to that, the possibility of causing allergen is very low. However allergenicity assessments need to be carried out before this delayed-ripening fruits go to the market.
Weediness is another potential risk is on environmental. The risk is on the escape of transgenic event to wild relatives of papaya, and converting them into an uncontrollable weed. But the possibility of GM delayed ripening papaya establishing weed is low and not likely to be greater than that of conventional papaya because papaya was never considered as a weed plant.
4. Transfer of Introduced Gene into Other Organisms
The next potential risk is about the transfer of the transgene into other organism (horizontal gene transfer). Horizontal gene transfer from GM delayed ripening papaya to non GM papaya is possible by pollination, except if buffer zone is not created. However the possibility of the introduced gene transferred to other organism is negligible. but even if such transfer occurred, it would be unlikely to pose any hazard to human health and safety or the environment, as the introduced genes are naturally present in papaya.
Beside the ACC oxidase gene, In my research a short sequences of DNA , CaMV 35 S promoter that are required for the expression ('switching on') are combine with ACC oxidase gene. It was reported that the introduction of viral DNA sequences into GM plants could produce new viruses through recombination ('gene exchange'). It was reported that there are natural barriers for this process to happen (Aaziz & Tepfer, 1999; Worobey & Holmes, 1999).There is only one reported case of recombination between plant and an animal virus Gibbs & Weiller, 1999). However there is no evidence saying that new viruses is created and eating infected plants can cause serious disease.
However, before this GM delayed ripening Eksotika papaya is ready for commercial planting and release, safety assessments (risk assessment and risk management will be done. The assessments will cover topics as follows:
The potential risk of transgenic papaya producing new kind of toxin
The potential of allergenic reaction to consumers
The nutritional content compared to the conventional counterparts
The general chemical; composition of the novel food compared to conventional counterparts
The evaluation of the methods used to develop the transgenic crop, including the molecular biological data which characterizes the genetic change