Team:Paris Bettencourt/Protocols/dneasy-blood-and-tissue-kit
Ferment It Yourself
iGEM Paris-Bettencourt 2O15
- Background
- Design
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Notebook
Vitamin A Vitamin B2 Vitamin B12 Phytase Riboswitch Differentiation on E. coli Differentiation on S. cerevisiae Manufacturing Idli and Micro-organisms Yeast DNA extraction using DNeasy® Blood & Tissue Kit
- Centrifuge a maximum of 5x106 cells for 5 minutes at 300g (190rpm). Resuspend in 200µL PBS. Add 20µL zymolyase.
- Add 3 volume of QG buffer (100mg of gel ~ 100ul) in the tube and incubate for 10 min at 50°C (or 10 min on a vortex).
- Add one volume of isopropanol and mix.
- vortex the tube for 2 min at 14 000 rpm.
- Carefully pipette the supernanant in a QIAquick column.
- Put the column in a 2ml microcentrifuge tube and centrifuge it for 1 min at 14 000 rpm.
- Throw the supernatant.
- Add 750 uL of PE buffer in the column and centrifuge for 1 min and discard flow-through.
- Centrifuge again for 2 min and put the column in a new 1.5uL microcentrifuge tube.
- Elute the DNA by putting 50uL of water in the middle of the column and let it stand for 2 min.
- Centrifugate for 2min at 10 000 rpm