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Development of transgenic soybeans encoding major peanut allergens Ara h 2 or Ara h 6 as a model to evaluate potential matrix effects relative to the risk assessment of GM crops
Evaluation of the potential risks of food allergy presented by any new GM crop is an important part of the regulatory review process required by most countries. To manage food safety, developers should ensure that they have avoided introducing a known allergen and potentially cross-reacting proteins, as well as have eliminated the risk that the recombinant protein becomes a de novo allergen. However, no studies have addressed the matrix effects between recipient crops and the gene donor organisms on the potential risk of allergenicity of the newly expressed proteins in GM crops. The overall goal of this dissertation is to investigate potential differences in matrix effects on the newly introduced protein by developing two model transgenic soybeans encoding major peanut allergens, Ara h 2 or Ara h 6. The 2S albumins of peanut were chosen as model transgenic proteins because peanut is one of the most important sources of food allergy in most geographical regions and these proteins are dominant allergens. Additionally, although soybeans and peanuts are both legumes and have overall similar food component characteristics, there are marked differences in the strength of food allergic reactions. Development of these GM soybean lines includes characterization of the recombinant proteins and ultimately they can be used for comparison of allergenicity with peanut and non-GM soybean. The first step in this evaluation required the development of two transgenic soybean lines using Agrobacterium mediated transformation. The second aim was to purify gram-level Ara h 2 from peanut and characterize it. The third aim was to purify and characterize recombinant Ara h 2 (rAra h 2) expressed in the GM soybean for use in comparisons with the natural Ara h 2 (nAra h 2) from peanut. A different post-translational proteolysis was revealed existing between soybean and peanut. However, the circular dichroism measurement demonstrated rAra h 2 and nAra h 2 shared very similar secondary and tertiary structures. This result indicates although different matrix effects might exist between recipient crops and gene donor organisms, there is little possibility to introduce an increased risk of allergenicity of recombinant proteins in this developed model of GM crops.
Luan, Fulei, "Development of transgenic soybeans encoding major peanut allergens Ara h 2 or Ara h 6 as a model to evaluate potential matrix effects relative to the risk assessment of GM crops" (2015). ETD collection for University of Nebraska - Lincoln. AAI3689952.