Genetically Modified Soybeans: Equal Allergenicity as their Wild Type Counterparts?
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Genetically Modified Soybeans: Equal Allergenicity as Natural Counterparts?
Genetically Modified Soybeans: Equal Allergenicity as their Wild Type Counterparts?Katie Van Den EindeNovember 24, 2009Advisor: Dr. Chastain1OverviewIntroduction: GM foods, allergies, controversyPaper 1Paper 2Paper 3Paper 4Current regulationsConclusions
2ImportanceGM foods:SoybeansCornTomatoesRiceCanolaPotatoesSugar beetsSugar cane
3ModificationsHerbicide resistanceInsect resistanceDisease resistanceAddition of proteins/vitamins
2003 84% of US soybean acreage was glyphosate tolerant (Roundup ready)
4Basics of Genetic ModificationProcedures1. Plasmid insertion2. Gene guns3. Protoplasts
5AllergiesMajority of allergic reactions are immunoglobulin E (IgE) mediated.IgE allergies affect about 1-2% of adults2-8% of children6Symptoms:Itchy, watery eyesRash CongestionItchinessDifficulty breathingAnaphylactic shock (Can be life threatening)
7Basics of allergic reactions1-Allergen2-IgE antibodies3-Mast cells4-Histamine release
8Anti-Histamines
9GM ControversyEthicsGene flowResistanceHarm to other organismsAllergens???
10OverviewIntroduction: GM foods, allergies, controversyPaper 1Paper 2Paper 3Paper 4Current regulationsConclusions
11Paper 1:Identification of a Brazil-nut allergen in transgenic soybeansNew England Journal of Medicine 1996-2S albumin to add methionine to soybeans because they dont have it12Purpose:To assess ability of proteins from
1)soybeans (Glycine max)2)transgenic soybeans3)Brazil nuts (Bertholletia excelsa)4)purified 2S albumin to bind to IgE serum13Methods:Radio allergosorbent test (RAST) 4 serumsSodium dodecyl sulfate-polyacrylamide-gel electrophoresis (SDS-PAGE) 9 serumsSkin Prick Tests
Serum from those allergice to brazil nuts14RAST basics
15Results: RAST
More inhibition of IgE binding = more allergic.Triangles= WTSquares= GM soybeanCircles= Brazil nutAdded human serum first, which bound if allergic. The more that bound, the more the labeled IgE binding was inhibited16Results: SDS-PAGE
IgE bindingTotal ProteinsIgE bindingStandardsWT SBTG SBBrazil Nut2SWT SBTG SBBrazil Nut2S2S in TG SBBrazil Nut17Results: Skin-Prick Test
Also on three control people with no allergies only had positive response to the histamine prick18Main 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.19OverviewIntroduction: GM foods, allergies, controversyPaper 1Paper 2Paper 3Paper 4Current regulationsConclusions
20Paper 2Lack of detectable allergenicity of transgenic maize and soya samplesJournal of Allergy and Clinical Immunology 200521Purpose:Monitor 5 GM products whose transgenes came from sources with no allergenic history
Such as bacteria22Methods:Food SurveyPrevious exposure?Skin Prick Tests27 kids with food allergies50 patients with asthma rhinitisSDS-PAGE23Flour products tested
24Food survey results
25Western Blot
26Testing Lab Supply
SDS PAGEWestern Blot27
SDS PAGEWestern Blot28Skin prick and IgE results
29Main Point:No detectable difference in IgE reactivity between wild type and GM soybeans or corn.
30OverviewIntroduction: GM foods, allergies, controversyPaper 1Paper 2Paper 3Paper 4Current regulationsConclusions
31Paper 3A comparative study of the allergenic potency of wild-type and glyphosate-tolerant gene-modified soybean cultivarsActa pathologica, microbiologica et immunologica Scandinavica 2003
32Purpose:To compare allergenicity of 8 wild type and 10 GM soybeans varieties (all for CP4 EPSPS)EPSPS: 5-Enolpyruvylshikimate-3-phosphate synthase33Methods:RAST (serum from 10 patients)SDS-PAGEHistamine Release testSkin prick tests34RAST results
More inhibition of labeled IgE binding = more original serum bound first.35RAST results
Concentration of extract needed for 50% inhibition of IgE binding (variety #12)If GM was more allergenic, the dots wouldve been way father down- hardly any needed, and the WT wouldve been really high up lots needed for the IgE binding36Histamine Release results
Notice lack of any major differences no where to point an arrow!Skin Prick TestHistamine Release(0=negative,6=lots)Also did statistical analysis of these found differences between the patients, but no significant difference between the two groups of soybeans37Histamine Release for patient I
Pretty similar!38Main Points:Difference between patients response, but no statistical difference between WT and TG soybeans.Addition of CP4 EPSPS gene higher allergenicity
39OverviewIntroduction: GM foods, allergies, controversyPaper 1Paper 2Paper 3Paper 4Current regulationsConclusions
40Paper 4Genetic modification removes an immunodominant allergen from soybeanPlant Physiology 200341Purpose:To silence the Gly m Bd 30K (P34) gene transgenically
42P34A major soybean allergenMore than 65% of soy-sensitive patients react only to the P34 proteinLess than 1% of total proteinPigs, calves and salmon also allergic
43Methods:Created a P34 silencing vector (plasmid pKS73)Grew these into homozyous strainsUsed SDS-PAGE for presence of P34 protein
44Results
Monoclonal antibodies45Soybean Protein Map
46Protein Analysis
Wild typeP34 SilencedMissing P34proteins and intermediates47Main Points:TG and WT were indistinguishable in size, shape, protein and oil content
P34 gene silencing was successful
48OverviewIntroduction: GM foods, allergies, controversyPaper 1Paper 2Paper 3Paper 4Current regulationsConclusions
49Whos in charge?Coordinated Framework for the Regulation of Biotechnology - 19863 regulatory bodiesof genetically modified foods:
(Animal and Plant Health Inspection Service)50Considerations:Effect on environment (animals, insects)Transferable to wild typeDigestive stabilityToxicityWeediness
51FDAFood additivesManufacturers responsible for checkingVoluntary consultation process - but all on U.S. market have undergone
52ConclusionsAllergens can be addedMostly, there is no differenceCan also remove allergensContinue studiesContinue monitoring
53Additional Works ConsultedUSDA Website. Biotechnology FAQs. Accessed 11/21/2009. http://www.usda.gov/wps/portal/!ut/p/_s.7_0_A/7_0_1OB?contentidonly=true&navid=AGRICULTURE&contentid=BiotechnologyFAQs.xml
Singer, S., Raven, D., Johnson G., Losos, J. 2005. Biology 7th Edition. McGraw Hill. New York, NY. 54Picture Referenceshttp://agriculture.sc.gov/UserFiles/Image/soybeans7.jpgstatistihttp://tharwacommunity.typepad.com/tharwa_review/images/2008/03/12/gm_foods.jpghttp://www.mun.ca/biology/desmid/brian/BIOL2060/BIOL2060-20/2032.jpghttp://repairstemcell.files.wordpress.com/2009/03/genetically-modified-food-fda.jpghttp://animalid.aphis.usda.gov/nais/wp_images/extension/high_res/usda-logo.jpghttp://blogs.venturacountystar.com/motorhead/epa.jpghttp://www.marlerblog.com/uploads/image/fda-logo.jpghttp://web.chemistry.gatech.edu/~williams/bCourse_Information/4581/techniques/gel_elect/gel.jpghttp://www.life.umd.edu/classroom/bsci423/songhttp://media.photobucket.com/image/Ige%20allergy/belldandy_84/Allergies.jpg/F03-44.jpghttp://api.ning.com/files/f7sw9nvb2lvWKi0Z-603fV67e5PN0http://www.flourallergy.com/images/allergy-test.jpg Y5iFz4Ef69JQNJKYzZ5lyynC5e9rpsiR7KJHFqW*CGRvzuPN6AianENPQ159UhHB680/pha0155l.jpghttp://www.worldcommunitycookbook.org/season/guide/photos/corn.jpghttp://e-internetbusiness.com/wp-content/uploads/2009/06/benadryl.png http://www.ars.usda.gov/is/graphics/photos/sep02/k9975-3i.jpghttp://intmedweb.wfubmc.edu/grand_rounds/1999/Image15.jpg55 Questions??
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