Editing TG1 Competent Cells (section)

== [[Category:Antibodies]][[Category:Protocols]]Protocol 2 ==



Centrifuge at 1200g, 15 min, 4°C.<br/>10. Resuspend the pellets with a 10-mL pipet in cold 10% glycerol to a total<br/>volume of 250 mL and transfer to one centrifuge bottle. Keep cells on ice while<br/>resuspending.<br/>11. Centrifuge at 1200g, 15 min, 4°C.<br/>12. Remove supernatant with a pipet. Be careful because the pellet is soft here.<br/>Resuspend the pellet with a 10-mL pipet in 10% glycerol to a total volume of<br/>50 mL. Keep cells on ice while resuspending.<br/>13. Centrifuge at 1200g, 15 min, 4°C.<br/>14. Remove supernatant with a pipet. Be careful because the pellet is very soft here.<br/>Resuspend the pellet with a 10-mL pipet in residual glycerol. Keep cells on ice<br/>while resuspending. The final volume should be 5–10 mL/L original culture.<br/>15. Aliquot 200 μL/cryotube and quick-freeze in liquid nitrogen (wear safety glasses<br/>when doing this). Store cells at –80°C.<br/>16. Transformation efficiency of cells should be determined by electroporating 100 pg<br/>control (uncut) vector DNA and plating onto appropriate selective medium (see<br/>Subheading 3.13.). Cells should yield 109 transformants/μg DNA.