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Team:UEA-JIC Norwich/Methods
From 2011.igem.org
High Efficiency Transformation Protocol
Culture Preparation Protocol
Glycerol Stock Solution Protocol
Qiagen Miniprep Protocol
Gel Electrophoresis Protocol
PCR protocol
PEG Transformation protocol
Algae Transformation using glass beads protocol
Algae Transformation using electroporation protocol
1. Transfer 50ml of algae culture into four 50ml screw top tubes.
2. Place all four tubes in the eppendorf centrifuge at 800rpm for 9 minutes at 20-22°C.
3. Immediately pour contents of tubes into glass beaker and keep tubes upside down so no unwanted suspension mixes with the pellet.
4. For all tubes resuspend cells in 250µl of TAP and sucrose media.
5. Add linearised G-Luc plasmid (5µL)to the suspension and place on ice.
6. Transfer 250µL of algae suspension into 1mL cuvette.
7. Subject cuvettes to electroporation at 2200v/ cm and place back on ice.
8. Add 500µL of starch solution to each cuvette and aspirate gently.
9. Add 750µL of each sample onto an LB plate.
10.Parafilm the lid and plate to seal and place in a light incubator.
Starch Preparation
1. Wash starch powder with 10mL distilled water and resuspend.
2. Centrifuge for five minutes at 4000rpm.
3. Pour off supernatant.
4. Resuspend with 500µL Ethanol.
5. Centrifuge for five minutes at 4000rpm.
6. Resuspend in 500µL TAP.
7. Centrifuge for five minutes at 4000rpm, pour off supernatant, and repeat once.
8. Resuspend in 500µL TAP with sucrose.
