Genetically Modified Organisms

1. Genetically Modified Organisms Image: www.ars.usda.gov

2. GMOs: Genetically Modified Organism Any organism that has had it’s genetic material changed or manipulated in some way, usually as a result of human intervention. Image: www.life.uiuc.edu Image: www.aggie-horticulture.tamu.edu This is the ancestor of what crop?

3. Strawberries with fish genes? Anti-freeze proteins in strawberry and tomato plants http://www.geo-pie.cornell.edu/media/fishberries.html

4. Trait targets of traditional plant breeding • Increase size (yield) of fruit or grain • Height of plants (taller OR shorter) • Flowering time to increase the number of crops in a year (rice) • Drought or cold tolerance • Appearance for ornamentals Image: www.news.cornell.edu Image: www.dlc.fi Image: http://faculty.etsu.edu/mcdowelt Image: www.jacksonandperkins.com Traditional breeding is limited to available genetic material in closely related species

5. Traits for genetically engineered organisms • Increase nutrient content (example: Golden Rice) • Delay ripening of tomatoes for better shipping (example: Flavr-savr) • Resistance to naturally occurring pests (example: Bt cotton) • Resistance to otherwise harmful herbicides • (example: RoundUp Ready soybeans) Image: www.scidev.net If there is a gene out there, it can be used, regardless of the source.

6. GMO Labelling • Currently, only two products on the market are “labeled” for altered nutrient content: • High laurate canola • high oleic soybean http://www.carleton.ca/catalyst/2005/s9.html

7. Agrobacterium tumefaciens: transfer of DNA into plants • Naturally occurring soil bacterium that causes crown gall disease • Bacteria contains the Ti (tumor-inducing) vector- a plasmid we can manipulate! Image: www.arabidopsis.info/students

8. Ti vector: modified for genetic engineering • Remove the tumor-inducing genes • Replace with gene of interest and promoter • Anything between the R and L border will be transferred.

9. Bt: example of genetic modification • Bacteria called Bacillus thuringiensis makes a protein with a crystal structure • The protein, when eaten by lepidoptera (caterpillars), sticks to the gut wall of the insect • Causes starvation and the dissolving of the internal organs • Purified protein has been used by organic farmers for years as a spray (is this a pesticide or herbicide?) Images: www.deh.gov.au

10. GMOs in the Market At least 70% of processed foods contain genetically modified ingredients: • Corn- in packaged foods (corn syrup, corn starch, etc.) • Cotton- Cotton isn’t just clothing: • cottonseed oil is present in many food items • Soy -in packaged foods and animal feed • Canola- the healthy oil

11. http://www.ers.usda.gov/data/biotechcrops/

12. Genetically modified foods: Others • Radicchio • Flax • Rice • Squash • Wheat • and more... • Papayas • Potatoes • Tomatoes • Sugarbeets • Cantaloupe • Banana Image: www.ebfarm.com Image: www.seedexseed.com While these have been modified and approved, you are most likely not eating these

13. Modifications currently approved • Herbicide tolerance (soy, cotton, corn, radicchio, sugarbeet, flax, rice, bentgrass, wheat, alfalfa) • Pesticide production (cotton, corn, potato, tomato) • Disease resistance (papaya, squash, potato, plum) • Delay in ripening (tomato, cantaloupe) • Improved (healthier) oil content (canola, soybean) • Reduced nicotine content (tobacco) • Increased amino acid content (corn)

14. On the SF bart… What do you think?

15. 8/07 NHM Genes for which we are testing: RuBisCo, Bt, 35S • RuBisCo- 599 bp- found only in plants. An enzyme that converts carbon dioxide into sugars and carbohydrates for the plant. This is our internal control. Most abundant protein found in nature. • Bt- 421 bp cry gene (toxin gene) will indicate that the plant is genetically modified • 35S promotor of the cauiliflower mosaic virus (CaMV 35s 221 bp -promoter element common in most transgenic plants. Its the “start” of a gene. Also indicates the plant is genetically modified)

16. Testing for GMOs in the field or pantry ELISA uses a color-change system to signal the presence of a protein. Limits: protein must not be denatured, as is often the case in processed foods. www.ilcrop.com PCR uses gene-specific primers to detect the presence of the transgene itself. Limits: expensive, not able to do in the field. * Enzyme-Linked ImmunoSorbent Assay

17. Staple food world-wide is deficient in Vit. A Ingo Potrykus, et al. developed rice that produces beta carotene, precursor to Vit. A Still not available for human consumption Golden Rice www.goldenrice.org

18. Papaya ring spot virus found in Hawaiian crops Cornell and Univ. Hawaii researchers developed papaya that produces viral coat protein Hawaiian papayas www.ctahr.hawaii.edu

19. DNA cassette PROMOTER GENE OF INTEREST SCREENABLE/ SELECTABLE MARKER

20. Genetically modified foods: Corn • Bt to protect from European corn borer • RoundUp Ready (or other herbicide tolerance), to allow spraying of herbicides • Stacked varieties Image: msucares.edu • Corn and corn derivatives are found in almost all packaged foods (corn syrup, corn starch, etc.) • In 2009, 85% of U.S. corn planted was genetically engineered

21. Genetically modified foods: Soy • Herbicide tolerance is the only modification of soy • Soy and derivatives are found in most packaged foods • Important as animal feed • In 2009, 91% of soy planted in the U.S. was engineered Image: cropwatch.unl.edu

22. Genetically modified foods: Cotton • Both Bt and herbicide tolerant varieties • Cotton isn’t just clothing: • cottonseed oil is present in many food items Image: ipm.ncsu.edu • In 2009, 88% of U.S. cotton planted was engineered

23. 3. There are 2 sequences of DNA that are most associated with GMO’s. • 35 S promotor of the cauiliflower mosaic virus (CaMV 35s) • Terminator of the nopaline synthase (NOS) gene of Agrobacterium tumefaciens. • We will look for either or both of these sequences in the food that we test. • PCR will allow us to isolate those segments of DNA and copy them. • An electrophoresis will give us a visual of the segments.

24. 8/07 NHM Genetically modified foods: Canola • Herbicide tolerant varieties • Canola is touted as one of the “healthiest” oils • In 2005, 80% of Canadian canola was engineered Image: canola-council.org

25. DNA sequence based on a known gene Host DNA Plasmid (circular DNA from bacteria) Electroporate into Agrobacterium tumefaciens + ligase and open plasmid gene of interest (goi) gene of interest selectable marker plasmid leaf disks Screen for transformants using selectable marker Incubate leaf disks with Agrobacterium Transfer to shoot and root growth media Each cell of the plant contains the new gene of interest Agrobacterium injects the gene of interest into the plant cell nucleus The gene of interest is inserted into the chromosomes of the plant with helper proteins Pot transformed plants and propagate

26. PCR is a method to amplify DNA Transfer genes by transformation- what do you need? A plasmid The bacteria Agrobacterium has a Ti plasmid (tumor inducing plasmid) that allows you to insert new genes (foreign DNA) regardless of size). Normally Ti causes tumors Scientist have engineered a cassette (sequence of genes) and are able to insert new genes much more easily. Bt gene confers resistance to insects

27. B. GMO PCR product Check plant pcr: is there a 455 bp band from GMO + (lane 5)? Check GMO PCR: is there a 203 bp band from GMO + (lane 6)

28. Types of genetic modification in plants • Selective breeding • selecting traits • marker assisted breeding • radiation/ mutagen induced • Hybridization (triticale) • Grafting (rootstock and scion) • Genetic engineering Image: farrer.csu.edu.au