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2. Cell transformation
From 2008.igem.org
(New page: 1. Remove vial of pre-prepared TOP10 chemically competent cells from -80˚C freezer on 5th floor then place in ice bucket to allow melting (will melt on ice). 2. Add between 10-100ng of DN...) |
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| - | 1. Remove vial of pre-prepared TOP10 chemically competent cells from -80˚C freezer on 5th floor then place in ice bucket to allow melting (will melt on ice). | + | 1. Remove vial of pre-prepared TOP10 chemically competent cells from -80˚C freezer on 5th floor then place in ice bucket to allow melting (will melt on ice). <br> |
| - | 2. Add between 10-100ng of DNA solution (if comes from DNA extraction, estimation may be necessary). | + | 2. Add between 10-100ng of DNA solution (if comes from DNA extraction, estimation may be necessary).<br> |
| - | 3. Mix the solution gently by tapping – anything more vigorous will result in potential damage to the cells. | + | 3. Mix the solution gently by tapping – anything more vigorous will result in potential damage to the cells.<br> |
| - | 4. Incubate on ice for 30 minutes, meanwhile ensure that a water bath is set to 42˚C and pre-heat a vial of SOC medium. | + | 4. Incubate on ice for 30 minutes, meanwhile ensure that a water bath is set to 42˚C and pre-heat a vial of SOC medium. <br> |
| - | 5. Once incubation on ice is completed, transfer the transformation tube to the 42˚C water bath for exactly 30 seconds. | + | 5. Once incubation on ice is completed, transfer the transformation tube to the 42˚C water bath for exactly 30 seconds.<br> |
| - | 6. Remove the transformation tube from the water bath and add 250µL of SOC medium, then rapidly place on ice for 1 min while being transported to a 37˚C incubator room with a shaker. | + | 6. Remove the transformation tube from the water bath and add 250µL of SOC medium, then rapidly place on ice for 1 min while being transported to a 37˚C incubator room with a shaker.<br> |
| - | 7. Shake at 37˚C for 1 hour. | + | 7. Shake at 37˚C for 1 hour.<br> |
8. Plate on appropriate media and antibiotics overnight, then check for colonies. | 8. Plate on appropriate media and antibiotics overnight, then check for colonies. | ||
| + | <br> | ||
Latest revision as of 20:16, 13 June 2008
1. Remove vial of pre-prepared TOP10 chemically competent cells from -80˚C freezer on 5th floor then place in ice bucket to allow melting (will melt on ice).
2. Add between 10-100ng of DNA solution (if comes from DNA extraction, estimation may be necessary).
3. Mix the solution gently by tapping – anything more vigorous will result in potential damage to the cells.
4. Incubate on ice for 30 minutes, meanwhile ensure that a water bath is set to 42˚C and pre-heat a vial of SOC medium.
5. Once incubation on ice is completed, transfer the transformation tube to the 42˚C water bath for exactly 30 seconds.
6. Remove the transformation tube from the water bath and add 250µL of SOC medium, then rapidly place on ice for 1 min while being transported to a 37˚C incubator room with a shaker.
7. Shake at 37˚C for 1 hour.
8. Plate on appropriate media and antibiotics overnight, then check for colonies.
