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Team:UQ-Australia/Notebook/Transformation procedure
From 2009.igem.org
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2. Gently mix cells, then aliquot 100ul competent cells into chilled polypropylene tubes. | 2. Gently mix cells, then aliquot 100ul competent cells into chilled polypropylene tubes. | ||
| - | 3. Add 1-10ng of plasmid DNA, moving | + | 3. Add 1-10ng of plasmid DNA, moving the pipette through cells while dispersing. Gently tap tubes to mix. |
4. Incubate cells on ice for 30 minutes | 4. Incubate cells on ice for 30 minutes | ||
Latest revision as of 02:16, 19 August 2009
Transformation Procedure
1. Thaw competent cells on ice. Place number of required polypropylene tubes on ice.
2. Gently mix cells, then aliquot 100ul competent cells into chilled polypropylene tubes.
3. Add 1-10ng of plasmid DNA, moving the pipette through cells while dispersing. Gently tap tubes to mix.
4. Incubate cells on ice for 30 minutes
5. Heat-shock cells for 30 seconds in 42'C waterbath. DO NOT SHAKE
6. Place on ice for 2 minutes
7. Add 900ul of room temperature SOC medium
8. Shake at 225rpm (37'C) for 1 hour
9. During the 1 hour incubation take agar plates out of fridge and incubate in 37'C incubator for 30 minutes
10. Spread 3 agar plates: 50ul, 100ul, 500ul
11. Incubate at 37'C overnight
