If the required gene has been isolated by shotgun approach, care must be taken to:
| 1. | cut the plasmid with a different restriction enzyme than that used to obtain the donor gene |
| 2. | cut the plasmid with the same restriction enzyme that was used to obtain the donor gene |
| 3. | avoid cutting the plasmid |
| 4. | cut the plasmid with two different restriction enzymes |
| Medium with Tetracycline | Medium with Ampicillin | |
| 1. | Growth | No growth |
| 2. | No growth | Growth |
| 3. | No growth | No growth |
| 4. | Growth | Growth |