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Team:British Columbia/Protocols/Beetle
From 2011.igem.org
Things to think about when designing your project and experiments, as well as general safety rules.
Site Directed Mutagenesis
A molecular biology technique in which a mutation is created at a defined site in a DNA molecule.
Bacterial Standard Operating Protocols
How to prepare competent cells, transform your construct into competent cells, and express your protein from a lac promoter.
Yeast Standard Operating Protocols
How to transform your construct into yeast, obtain crude extract for SDS-PAGE, and perform GFP fixation for microscopy and fluorescence-activated cell sorting.
Gas Chromatography-Mass Spectrometry (GC-MS)
A method that combines the features of gas-liquid chromatography and mass spectrometry to identify different substances within a test sample.
Beetle Transfer Experiments
Preliminary experiments to probe the efficiency of transferring yeast via beetle vector.
Yeast-Fungi Co-culture Experiments
Preliminary experiments to investigate the competitive interactions between yeast and the Bluestain Fungus.
Beetle Transfer Experiment
In this experiment, we wanted to find out whether the beetles could serve as a vector for our genetically modified yeast. To do this, we followed the following procedure.
1) Obtain beetles
2) Place beetles using small sterile forceps on plates with appropriately labeled yeast (GFP or Clonat resistance). Allow beetles to walk on the yeast for 1 minute.
3) Place the beetles on an empty plate for various amounts of time (
4) Place beetles on new, clean YPD plates. Allow the beetle to walk on the clean plate for one minute.
5) Remove and properly dispose of the beetle. Allow the plate to grow for 2-3 days.
6) All plates should have yeast colonies, even the control. Streak out the colonies on appropriate plates (Note auxotrophy and antibiotic resistance, although the former is not diagnostic). Allow to grow 1-2 days.
7) Check the plates. The control should not have grown and the clonat plates should have colonies only from the appropriate plates. Follow the protocol for GFP fixation and FACS onthis page to check for GFP activity.
