Joyce:Transformation

Transformation

Procedure

1. Obtain an ependorff of competent cells
2. Place the cells on ice and let them thaw on ice (about 10-15 minutes)
3. Gently stir the cells with a pipette tip to resuspend them
4. Pipette 50 ul of cells into an ependorff
5. With gentle stirring, pippette 5 ul of a 20 ul ligation reaction into the ependorff
6. Place the ependorff on ice for at least 30 minutes
7. Prepare the heating block for 42*C
8. Heat shock the transformation mixture for 45 seconds at 42*C
9. Put the transformation mixture on ice for 2 minutes
10. Add 400 ul of sterile YT to the transformation mixture
11. Mix gently by inverting several times
12. Incubate at 37*C for at least 45 minutes
13. Centrifuge at full speed for 2 minutes
14. Discard 300 ul of the supernatant
15. Resuspend the pellet in the remaining 150 ul
16. Plate