"
Virginia Commonwealth/27 May 2009
From 2009.igem.org
|
|
|
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Wednesday 27 May 2009
Tasks
- Transform NEB10beta with J04450 and I13521, using frozen stock of comp. cells.
Results
- Kept all tubes and cuvettes on ice at all times, and thawed one tube of NEB10beta competent cells (250uL). Each of the following transformation was performed individually:
- (-) control, with no DNA added
- 1uL J04450 and 50uL cells
- 2uL J04450 and 50uL cells
- 1uL I13521 and 50uL cells
- Added 50uL cells to cold microcentrifuge tube.
- To cells, added appropriate volume and type of DNA, mixing well by pipette, and returning to ice to incubate for 1min.
- Transfer mix to cold 2mm electrotransformation cuvette, tapping to get mixture to the bottom.
- Pulsed cuvette on Ec2 setting, then immediately added 1mL LB.
- Transferred mixture to 15ml centrifuge tube and incubated for 30min at 37*C.
- Spread 35uL on either kan (J04450 and ctrl) or amp (I13521), incubated at 37*C overnight.
