Genetically Modified Soybeans: Equal Allergenicity as their Wild Type Counterparts? Katie Van Den Einde November 24, 2009 Advisor: Dr. Chastain
Overview Introduction: GM foods, allergies, controversy Paper 1 Paper 2 Paper 3 Paper 4 Current regulations Conclusions
Importance GM foods: ◦ Soybeans ◦ Corn ◦ Tomatoes ◦ Rice ◦ Canola ◦ Potatoes ◦ Sugar beets ◦ Sugar cane
Modifications Herbicide resistance Insect resistance Disease resistance Addition of proteins/vitamins 2003 – 84% of US soybean acreage was glyphosate tolerant (Roundup ® ready)
Basics of Genetic Modification Procedures ◦ 1. Plasmid insertion ◦ 2. Gene “guns” ◦ 3. Protoplasts
Allergies Majority of allergic reactions are immunoglobulin E (IgE) mediated. IgE allergies affect about 1-2% of adults 2-8% of children
Symptoms: Itchy, watery eyes Rash Congestion Itchiness Difficulty breathing Anaphylactic shock (Can be life threatening)
Basics of allergic reactions 1-Allergen 2-IgE antibodies 3-Mast cells 4-Histamine release
Anti-Histamines
GM Controversy Ethics Gene flow Resistance Harm to other organisms Allergens???
Overview Introduction: GM foods, allergies, controversy Paper 1 Paper 2 Paper 3 Paper 4 Current regulations Conclusions
Paper 1: Identification of a Brazil-nut allergen in transgenic soybeans ◦ New England Journal of Medicine 1996
Purpose: To assess ability of proteins from 1)soybeans (Glycine max) 2)transgenic soybeans 3)Brazil nuts (Bertholletia excelsa) 4)purified 2S albumin to bind to IgE serum
Methods: Radio allergosorbent test (RAST) – 4 serums Sodium dodecyl sulfate-polyacrylamide-gel electrophoresis (SDS-PAGE) – 9 serums Skin Prick Tests
RAST basics
Results: RAST More inhibition of IgE binding = more allergic. Triangles= WT Squares= GM soybean Circles= Brazil nut
Results: SDS-PAGE IgE binding Total Proteins IgE binding
Results: Skin-Prick Test
Main Points: GM soybean protein successfully competed with Brazil nut protein. IgE from 8/9 allergic to Brazil nut bound to introduced 2S albumin in GM soybeans.
Overview Introduction: GM foods, allergies, controversy Paper 1 Paper 2 Paper 3 Paper 4 Current regulations Conclusions
Paper 2 Lack of detectable allergenicity of transgenic maize and soya samples Journal of Allergy and Clinical Immunology 2005
Purpose: Monitor 5 GM products whose transgenes came from sources with no allergenic history
Methods: Food Survey ◦ Previous exposure? Skin Prick Tests ◦ 27 kids with food allergies ◦ 50 patients with asthma rhinitis SDS-PAGE
Flour products tested
Food survey results
Western Blot
Testing Lab Supply SDS PAGE Western Blot
SDS PAGE Western Blot
Skin prick and IgE results
Main Point: No detectable difference in IgE reactivity between wild type and GM soybeans or corn.
Overview Introduction: GM foods, allergies, controversy Paper 1 Paper 2 Paper 3 Paper 4 Current regulations Conclusions
Paper 3 A comparative study of the allergenic potency of wild-type and glyphosate- tolerant gene-modified soybean cultivars ◦ Acta pathologica, microbiologica et immunologica Scandinavica 2003
Purpose: To compare allergenicity of 8 wild type and 10 GM soybeans varieties (all for CP4 EPSPS)
Methods: RAST (serum from 10 patients) SDS-PAGE Histamine Release test Skin prick tests
RAST results More inhibition of labeled IgE binding = more original serum bound first.
RAST results Concentration of extract needed for 50% inhibition of IgE binding (variety #12)
Histamine Release results Notice lack of any major differences – no where to point an arrow! Skin Prick Test Histamine Release (0=negative, 6=lots)
Histamine Release for patient I Pretty similar!
Main Points: Difference between patients’ response, but no statistical difference between WT and TG soybeans. Addition of CP4 EPSPS gene ≠ higher allergenicity
Overview Introduction: GM foods, allergies, controversy Paper 1 Paper 2 Paper 3 Paper 4 Current regulations Conclusions
Paper 4 Genetic modification removes an immunodominant allergen from soybean ◦ Plant Physiology 2003
Purpose: To silence the Gly m Bd 30K (P34) gene transgenically
P34 A major soybean allergen More than 65% of soy-sensitive patients react only to the P34 protein Less than 1% of total protein Pigs, calves and salmon also allergic
Methods: Created a P34 silencing vector (plasmid pKS73) Grew these into homozyous strains Used SDS-PAGE for presence of P34 protein
Results
Soybean Protein “Map”
Protein Analysis Wild typeP34 Silenced Missing P34 proteins and intermediates
Main Points: TG and WT were indistinguishable in size, shape, protein and oil content P34 gene silencing was successful
Overview Introduction: GM foods, allergies, controversy Paper 1 Paper 2 Paper 3 Paper 4 Current regulations Conclusions
Who’s in charge? Coordinated Framework for the Regulation of Biotechnology regulatory bodies of genetically modified foods: (Animal and Plant Health Inspection Service)
Considerations: Effect on environment (animals, insects…) Transferable to wild type Digestive stability Toxicity “Weediness”
FDA Food additives Manufacturers responsible for checking Voluntary consultation process - but all on U.S. market have undergone
Conclusions Allergens can be added Mostly, there is no difference Can also remove allergens Continue studies Continue monitoring
Additional Works Consulted USDA Website. “Biotechnology FAQs.” Accessed 11/21/ d=AGRICULTURE&contentid=BiotechnologyFAQs.xml d=AGRICULTURE&contentid=BiotechnologyFAQs.xml Singer, S., Raven, D., Johnson G., Losos, J Biology 7 th Edition. McGraw Hill. New York, NY.
Picture References statistihttp://tharwacommunity.typepad.com/tharwa_review/images/2008/03/12/gm_foods.jpg Y5iFz4Ef69JQNJKYzZ5lyynC5e9rpsiR7KJHFqW*CGRvzuPN6AianENPQ159UhHB680/pha0155l.jpg
Questions??