1. Practical training: Proof of a gentechnical variation of foods with PCR

2. Principle preperation of a PCR 10`, 72 °C Completing 60``, 94 ° Denaturation 40 cycles 120``, 94°C Denaturation 60``, 55°C Annealing 120``, 72 °C Elongation

3. I. Extraction of DNA

4. 1. Preparation 1.1 Label 2 screw cap tubes on the top with: - non-GMO (- GMO) - food sample (T) and your group number! 1.2 Pipet 500 µl of homo- genised InstaGene- Matrix into each tube! There have to be the same amount of beads in both tubes! supernatant beads

5. 2. Extraction of DNA from „non- GMO food“ 2.1Weigh out 1 g of certified non- GMO food and put it into the mortar! 2.2Add 5 ml of distilled H 2 O!

6. 2.3 Homogenise the mixture for about five minutes! 2.4 Add 5 ml of H 2 O dist. and pestle until smooth enough to pipet the mixture! 2. Extraction of DNA from „non- GMO food“

7. 2.5 Pipet 25 µl of the food mixture into the liquid of the screw cap tube labeled “-GMO”! 2.6 Recap tube and shake well by flicking the tube! To avoid contamination with „+GMO DNA“ „-GMO material“ should be extracted first! Mortar and pestle should be cleaned with a detergent and rinsed out with distilled H 2 O!

8. 3. Extraction of DNA from food sample 3.1Weigh out 1 g of food sample to be tested. (T) and place it in the mortar! 3.2 Add 5 ml of H 2 O dist. ! Please use gloves!!!

9. 3.3Homogenise the mixture for about five minutes! 3.4Again add 5 ml of H 2 O dist. and pestle until smooth enough to pipet the mixture! 3. Extraction of DNA from food sample

10. 3.5Pipet 25 µl of the food mixture into the liquid of the screw cap tube labeled “T”! 3.6Recap tube and shake well by flicking the tube!

11. 4. Denaturation of enzymes at 95°C 4.1Place both screw cap tubes in water bath of 95 o C for 5 minutes! 4.2Centrifuge the tubes for 5 minutes at 13000 rpm! 4.3Put 50 µl of supernatant into two new tubes which have been labeled before!

12. Pay attention: Only take the supernatant without the InstaGene beads by removing of the food control DNA and the test food DNA!

13. II. Preparation: PCR and electrophoresis in group work According to their assignment, each group prepares a part of the PCR and the electrophoresis for all the other groups.

14. III. Preparation and carrying out of PCR

15. 1. Preparation of PCR-tubes 1.1 Number your PCR-tubes 1-6! 1.2 Pipet the PCR-preparations according to the following table! 1.3 Mix your PCR-preparations by pipeting up and down! 1.4 Cool your PCR-preparations on ice until starting PCR!

16. NrMaster MixDNA 110 µl PMM10 µl -GMO food control DNA 210 µl GMM10 µl -GMO food control DNA 310 µl PMM10 µl Test food DNA (T) 410 µl GMM10 µl Test food DNA (T) 510 µl PMM10 µl +GMO control DNA 610 µl GMM10 µl +GMO control DNA

17. Carry out the PCR according to the temperature program of the thermocycler! Pay attention on the position of your samples in the thermocycler to avoid confusion! Make sure that your tubes are correctly closed! 2. Carrying out of PCR Cycler- Program Overview of the tubes in the cycler

18. IV. Evidence of PCR-products by electrophoresis (Agarose/PAGE) 1.According to your assignment give evidence of the PCR products by carrying out Agarose- or Polyacrylamite-gel- electrophoresis! 2. According to your assignment dye your gels and analyse them! Filling the gelslots