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Team:Montreal/Cell transformation
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(New page: 1. Remove vial of pre-prepared TOP10 chemically competent cells from -80˚C freezer and thaw in ice. 2. Add between 10-100ng of DNA in solution (varies with cell competency). 3. Mix the ...) |
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| + | {|style="font color="#ffffff"; background-color:#cd0000; cellpadding="3" cellspacing="5" border="2" bordercolor="#cd0000"border-spacing:6px; text-align:center" width="960px" | ||
| + | !style="text-align:center; background-color:#cd0000; border-width:0px; padding:3px;"|[[Team:Montreal|<font color="#ffffff">Home</font>]] | ||
| + | !style="text-align:center; background-color:#cd0000; border-width:0px; padding:3px;"|[[Team:Montreal/Team|<font color="#ffffff">The Team</font>]] | ||
| + | !style="text-align:center; background-color:#cd0000; border-width:0px; padding:3px;"|[[Team:Montreal/Project|<font color="#ffffff">The Project</font>]] | ||
| + | !style="text-align:center; background-color:#cd0000; border-width:0px; padding:3px;"|[[Team:Montreal/Parts|<font color="#ffffff">Parts Submitted to the Registry</font>]] | ||
| + | !style="text-align:center; background-color:#cd0000; border-width:0px; padding:3px;"|[[Team:Montreal/Modeling|<font color="#ffffff">Modeling</font>]] | ||
| + | !style="text-align:center; background-color:#cd0000; border-width:0px; padding:3px;"|[[Team:Montreal/Notebook|<font color="#ffffff">Notebook</font>]] | ||
| + | |} | ||
| + | ==Protocol== | ||
| + | |||
1. Remove vial of pre-prepared TOP10 chemically competent cells from -80˚C freezer and thaw in ice. | 1. Remove vial of pre-prepared TOP10 chemically competent cells from -80˚C freezer and thaw in ice. | ||
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3. Mix the solution gently by tapping – anything more vigorous will result in potential damage to the competent cells. | 3. Mix the solution gently by tapping – anything more vigorous will result in potential damage to the competent cells. | ||
| - | 4. Incubate on ice for 30 minutes | + | 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. '''Heat shock''' the transformation tube | + | 5. '''Heat shock''' the transformation tube in a 42˚C water bath for exactly 30s. |
| - | 6. Remove the transformation tube from the water bath and add 250µL of SOC medium, then | + | 6. Remove the transformation tube from the water bath and add 250µL of SOC medium, then place on ice for 60s. |
7. Shake and incubate at 37˚C for one hour. | 7. Shake and incubate at 37˚C for one hour. | ||
8. Plate on appropriate media and antibiotics for approximately 20 hours, then check for colonies | 8. Plate on appropriate media and antibiotics for approximately 20 hours, then check for colonies | ||
Latest revision as of 19:41, 17 June 2008
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Protocol
1. Remove vial of pre-prepared TOP10 chemically competent cells from -80˚C freezer and thaw in ice.
2. Add between 10-100ng of DNA in solution (varies with cell competency).
3. Mix the solution gently by tapping – anything more vigorous will result in potential damage to the competent 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. Heat shock the transformation tube in a 42˚C water bath for exactly 30s.
6. Remove the transformation tube from the water bath and add 250µL of SOC medium, then place on ice for 60s.
7. Shake and incubate at 37˚C for one hour.
8. Plate on appropriate media and antibiotics for approximately 20 hours, then check for colonies
