Team:Groningen/Notebook/14 July 2009
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'''Plasmid Purification''' | '''Plasmid Purification''' | ||
- | Plasmid isolation was performed on the cultures of GVP and Terminator containing cells with the "Sygma-Aldrich™ GenElute™ Plasmid Miniprep Kit". | + | Plasmid isolation was performed on the cultures of GVP and Terminator containing cells with the "Sygma-Aldrich™ [http://www.sigmaaldrich.com/life-science/molecular-biology/dna-and-rna-purification/plasmid-miniprep-kit.html GenElute™] Plasmid Miniprep Kit". |
* From each tube 5mL of culture was collected in a 2mL cup, and the cells were pelleted by centrifugation for 1 min. at max. speed and the supernatant discarded. | * From each tube 5mL of culture was collected in a 2mL cup, and the cells were pelleted by centrifugation for 1 min. at max. speed and the supernatant discarded. | ||
* Cells were resuspended in 200μL Resuspension Solution by up and down pipetting. | * Cells were resuspended in 200μL Resuspension Solution by up and down pipetting. |
Revision as of 09:40, 14 July 2009
Wet
GVP Cluster
Plasmid Purification
Plasmid isolation was performed on the cultures of GVP and Terminator containing cells with the "Sygma-Aldrich™ GenElute™ Plasmid Miniprep Kit".
- From each tube 5mL of culture was collected in a 2mL cup, and the cells were pelleted by centrifugation for 1 min. at max. speed and the supernatant discarded.
- Cells were resuspended in 200μL Resuspension Solution by up and down pipetting.
- To the mixture 200μL Lysis Solution was added, mixed by inverting the cup, and stored at room temperature for 5 minutes.
- 350μL of Neutralisation Solution was added and the tubes inverted.
- Cell debri was pelleted by centrifugation at full speed for 10 min.
- Column was prepared by adding 500μL Column Preparation Solution and centrifuging for 1 min. full speed
- The lysate was transfered to the column and centrifuged for 1 min. full speed
- Column was washed with both washing solutions
- Plasmids were eluted with 30μL MQ and stored in the fridge
Concentration of Plasmids
The concentration of isolated plasmid was determined with the use of a nano-drop.
GVP eluted in MQ
- 329.7 ng/μL
- 1.84 (260/280)
- 2.13 (260/230)
BBa_J23109 eluted in MQ
- 174.9 ng/μL
- 1.88 (260/280)
- 1.92 (260/230)
BBa_J23100 eluted in MQ
- 155.3 ng/μL
- 1.85 (260/280)
- 1.79 (260/230)
BBa_J23106 eluted in MQ
- 219.4 ng/μL
- 1.86 (260/280)
- 1.93 (260/230)
Transporters
Dry
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