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Recovery

1.       Obtain an LB agar plate with the appropriate antibiotic.

2.       Using a sterile pipette tip, touch the bacteria growing within the punctured area of the stab culture. (A sterilized wire loop or sterile toothpick can be used in place of a sterile pipette tip.)

3.       Run this tip lightly over a section of the plate, as shown in the figure, to create streak #1.

4.       Using another sterile pipette tip, pass through streak #1 and spread the bacteria over a second section of the plate, to create streak #2.

5.       Using a third sterile pipette tip, pass through streak #2 and spread the bacteria over the last section of the plate, to create streak #3.

6.       Grow overnight in a 37 ^o C incubator (unless a different growth temperature is indicated on the plasmid datasheet).

7.       In the morning, single colonies should be visible. If the bacterial growth is too dense, re-streak onto a new agar plate to obtain single colonies.