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Protocol DNA extraction from blood samples (DOC)

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					Protocol: DNA extraction from blood samples

1. Transfer 80l of blood sample to a new eppendorf. 2. Add 1ml of NH4Cl to each tube, mix vigorously by inverting the tube and incubate at room temperature for 10 minutes. 3. Centrifuge for 5 minutes at 2000rpm at 4ºC. 4. Remove the supernatant. 5. Add 1ml of PBS and mix. 6. Centrifuge for 5 minutes at 2000rpm at 4ºC. 7. Remove PBS. 8. Ressuspend the pellet in the remaing PBS (50l). 9. Make a solution mix of lysis buffer (3ml) and proteinase K (3l). 10. Add 200l of the solution mix to each tube. 11. Incubate at 50ºC with mix (300rpm) during many hours. 12. Add 200l of Phenol:Chloroform Isoamyl Alcohol to each tube and mix vigorously by inverting the tube. 13. Centrifuge at maximum speed (13200) for 3 minutes. 14. Transfer the aqueous fase (top) to a new tube. 15. Add 200l of Isopropanol and mix by inverting the tube. 16. Centrifuge at 13200 rpm for 20 minutes. 17. Remove the supernatant. 18. Add 1ml of alcohol 70%. 19. Centrifuge at 13200 rpm for 10 minutes. 20. Remove the supernatant. 21. Dry the pellet at room temperature. 22. Add 20l of TE buffer to each tube. 23. Incubate at room temperature for 4h. 24. Determine concentration.


				
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