Team:Colombia Uniandes/Protocols

Protocol for extraction of yeast genome:

GenElute DNA Kit from Sigma-Aldrich with modifications

1. 1,5 mL 2 min x 15000 rpm.
2. 100 µL zymolyase solution.
3. Vortex.
4. Incubate 37°C x 30 min.
5. 180 µL of Lysis Solution T.
6. 20 µL of Proteinase K.
7. Incubate 37°C x 30 min.
8. 200 µL of Lysis Solution C.
9. Vortex x 15 s.
10. Incubate 55°C x 10 min.
11. Column Preperation --> 500 µL Column Preparation Solution --> 13000 rpm x 1 min – Discard the eluate.
12. 200 µL of ethanol (100%) to the lysate – Vortex. --> Transfer the entire contents of the tube into the binding column --> 12000 rpm x 1 min --> Discard the collection tube and place the column in a new one.
13. 500 µL of Wash Solution 1 to the column. --> 12000 rpm x 1 min --> Discard the collection tube and place the column in a new one.
14. 500 µL of Wash Solution Concentrate (+ ethanol) --> 15000 rpm x 3 min – Discard the collection tube and place the column in a new one.
15. 30 µL of water (miliQ) --> 14000 rpm x 1 min.
16. 30 µL of water (miliQ)  14000 rpm x 1 min.
17. The final eluate contains the DNA – Do not discard!