Bio-Rad Ligation and Transformation Module User Manual

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6. After bacteria have grown in C-growth medium for 20–40 min at 37°C

with shaking, transfer the culture to your competent cells tube by
decanting or pipetting it. It is better not to put the actively growing cell
culture on ice at this step.

7. Centrifuge the culture in a microcentrifuge at top speed for 1 min. Make

sure that the microcentrifuge is balanced and accommodate tubes of
classmates to ensure economic use of the microcentrifuge. Immediately
put the pelleted culture on ice.

Note: After this step, it is very important to keep the bacteria on ice as
much as possible during this procedure. Transformation efficiency will be
severely compromised if the cells warm up.

Note: It is very important to treat the bacteria extremely gently during this
procedure — the bacteria are very fragile and your transformation efficiency
will be compromised unless you are very gentle.

8. Locate the pellet of bacteria at the bottom of the tube. Remove the

culture supernatant, avoiding the pellet, using a 1,000 µl pipet or a
vacuum source. Keep the cells on ice.