Huang: DH5alpha transformation
From OpenWetWare
(Difference between revisions)
(New page: 1. Set tubes of frozen cells on ice and let thaw very slowly 2. Add 100ul thawed cells to new eppendorf 3. Add 1-2ul DNA to cells 4. Incubate on ice for 20 min 5. Heat shock in 42C water b...) |
Current revision (19:49, 1 April 2009) (view source) |
||
| Line 1: | Line 1: | ||
| - | 1. Set tubes of frozen cells on ice and let thaw very slowly | + | ==Overview== |
| - | 2. Add 100ul thawed cells to new eppendorf | + | |
| - | 3. Add 1-2ul DNA to cells | + | This protocol allows competent cell transformation. |
| - | 4. Incubate on ice for 20 min | + | |
| - | 5. Heat shock in 42C water bath for one min (make sure water bath is at exactly 42C) | + | ==Materials== |
| - | 6. Incubate on ice 2 min | + | *DH5alpha competent cells |
| - | 7. Add 900ul growth medium (LB) and shake 1 hour at 37C | + | *LB |
| - | 8. Plate about 200ul and grow overnight | + | *LB + antibiotic plates |
| + | *Plasmid DNA or DNA to transform the cells with | ||
| + | |||
| + | ==Procedure== | ||
| + | *1. Set tubes of frozen cells on ice and let thaw very slowly | ||
| + | *2. Add 100ul thawed cells to new eppendorf | ||
| + | *3. Add 1-2ul DNA to cells | ||
| + | *4. Incubate on ice for 20 min | ||
| + | *5. Heat shock in 42C water bath for one min (make sure water bath is at exactly 42C) | ||
| + | *6. Incubate on ice 2 min | ||
| + | *7. Add 900ul growth medium (LB) and shake 1 hour at 37C | ||
| + | *8. Plate about 200ul and grow overnight | ||
| + | |||
| + | |||
| + | ==Contact== | ||
| + | *[[User:Carolina Tropini|Carolina Tropini]] | ||
Current revision
Contents |
Overview
This protocol allows competent cell transformation.
Materials
- DH5alpha competent cells
- LB
- LB + antibiotic plates
- Plasmid DNA or DNA to transform the cells with
Procedure
- 1. Set tubes of frozen cells on ice and let thaw very slowly
- 2. Add 100ul thawed cells to new eppendorf
- 3. Add 1-2ul DNA to cells
- 4. Incubate on ice for 20 min
- 5. Heat shock in 42C water bath for one min (make sure water bath is at exactly 42C)
- 6. Incubate on ice 2 min
- 7. Add 900ul growth medium (LB) and shake 1 hour at 37C
- 8. Plate about 200ul and grow overnight
