Melbourne/Transformation Protocol
From 2007.igem.org
(Difference between revisions)
(→References) |
|||
Line 2: | Line 2: | ||
*Applications: | *Applications: | ||
*#Amplification of Biobrick DNA for storage and use. | *#Amplification of Biobrick DNA for storage and use. | ||
- | *#Selection | + | *#Selection and amplification of ligated constructs. |
*Time to complete protocol: | *Time to complete protocol: | ||
Line 11: | Line 11: | ||
====Method from primary and secondary reagents==== | ====Method from primary and secondary reagents==== | ||
=====Primary & secondary Reagents Required including controls===== | =====Primary & secondary Reagents Required including controls===== | ||
- | *Competent cells | + | *Competent cells (from -70degree freezer) |
*DNA for transformation | *DNA for transformation | ||
- | *LB | + | *LB (cupboard) |
- | *LB-agar plates with selective antibiotic | + | *LB-agar plates with selective antibiotic (cool room) |
=====Method including controls===== | =====Method including controls===== | ||
Line 32: | Line 32: | ||
*Ice box | *Ice box | ||
*Pipettes | *Pipettes | ||
- | *42 degree water bath | + | *42 degree water bath (balance room) |
- | *37 degree incubator | + | *37 degree incubator |
*Bunsen burner | *Bunsen burner | ||
*Spreader | *Spreader |
Revision as of 05:30, 2 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.
- Incubate at 37degrees 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