Melbourne/Transformation Protocol
From 2007.igem.org
(Difference between revisions)
m (Melb:Transformation Protocol moved to Melbourne/Transformation Protocol) |
(→Method including controls) |
||
Line 21: | Line 21: | ||
#Heat shock in water bath at 42 degrees for 1min. | #Heat shock in water bath at 42 degrees for 1min. | ||
#Incubate on ice for 15min. | #Incubate on ice for 15min. | ||
- | #Add 1mL LB. | + | #Add 1mL LB (flame tip before use). |
- | #Incubate at | + | #Incubate at 37 degrees for 1 hour. |
#Spin down cells and remove majority of LB. | #Spin down cells and remove majority of LB. | ||
#Resuspend cells in remaining LB. | #Resuspend cells in remaining LB. | ||
Line 28: | Line 28: | ||
#Incubate plate overnight at 37 degrees. | #Incubate plate overnight at 37 degrees. | ||
#Place in cold room until needed. | #Place in cold room until needed. | ||
+ | |||
=====Equipement Required===== | =====Equipement Required===== | ||
*1.5mL Microfuge tubes | *1.5mL Microfuge tubes |
Revision as of 06:22, 4 July 2007
<Return to list of protocols> <Team home page>
- Applications:
- Amplification of Biobrick DNA for storage and use.
- Selection and amplification of ligated constructs.
- Time to complete protocol:
- Lab time: 10min, 10min, 10min, 15min.
- Waiting time: 45min, 15min, 1hour, overnight.
- Approximate cost of materials: $
Method from primary and secondary reagents
Primary & secondary Reagents Required including controls
- Competent cells (from -70degree freezer)
- DNA for transformation
- LB (cupboard)
- LB-agar plates with selective antibiotic (cool room)
Method including controls
- Add 1uL resuspended plasmid DNA to 50uL competent cells.
- Incubate on ice for 45min.
- Heat shock in water bath at 42 degrees for 1min.
- Incubate on ice for 15min.
- Add 1mL LB (flame tip before use).
- Incubate at 37 degrees for 1 hour.
- Spin down cells and remove majority of LB.
- Resuspend cells in remaining LB.
- Under a bunsen spread resuspended bacteria on agar plate on selective antibiotic.
- Incubate plate overnight at 37 degrees.
- Place in cold room until needed.
Equipement Required
- 1.5mL Microfuge tubes
- Ice box
- Pipettes
- 42 degree water bath (balance room)
- 37 degree incubator
- Bunsen burner
- Spreader
References