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!

Glucocorticoid Sensor

Animals release glucocorticoids under stress situations such as famine, injuries, anxiety or fear. This hormone is the one we will detect as an indicator of stressful conditions for the animal. This differential equation expresses the concentration of glucocorticoids outside the cell, which only depends on their diffusion rate into the cell.






18. GRI: The amount of glucocorticoids inside the yeast cell depends on the degradation rate of the hormone, the complex formation and the import rate of the glucocorticoids inside the cell, which depends on the diffusion rate of the hormone in relation with the concentration of the hormone outside and inside the cell. </div> </div>