Team:Dundee/Team -




iGEM Sumbawagen 2014 · Econey

Notebook – Protocol – 3. Electrophoresis and DNA visualization

1.50x TAE (Thermoscientific) was diluted to 1x TAE (1 ml 50x TAE + 49 ml distilled water).

2. 3 g of Agarose S (Nippongene) was added into 200 ml of 1x TAE to get 1.5 % agarose gel in a glass flask.

3. Agarose was melted by putting the glass flask in boiled water – we have no microwave – and sometimes stirred slowly to avoid bubble formation with stirrer bar on a stirrer.

4. Completely melted agarose solution was poured into the casting tray with comb.

5. Let the gel became solid by incubating at room temperature for 30 minutes.

6. Electrophoresis was done in 1x TAE buffer in an electrophoresis tank (Mupid)

7. The volume of samples used for electrophoresis was 7 ul for 1 kB DNA ladder and 10 ul (PCR product), or 15 ul (Restriction enzyme digestion product) for DNA samples for each well.

8. Electrophoresis was done for 20 minutes using 135V setting.

9. After electrophoresis, gel was put into 1x TAE solution containing Sybrsafe to stain the DNA for about 20 minutes.

10. Then gel was placed on trans illuminator for viewing and documentation using pocket digital camera.

11. The file was viewed with computer using Photoshop.

12. For disposal, gel and buffer were dried then burnt together with any tissues or papers in contact.