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Team:UNIPV-Pavia/Methods Materials/XGal
From 2009.igem.org
- LB medium
- DNA resuspension from iGEM 2009 plates
- Transformation
- X-Gal staining protocol for beta galactosidase (blue/white screening)
- Glycerol stocks
- Plasmid extraction
- BioBrick digestion with restriction enzymes
- Electrophoresis
- DNA gel extraction
- DNA precipitation with sodium acetate
- Ligation
- PCR
- Sensing ethanol concentration: our do-it-yourself kit
X-Gal staining protocol for beta galactosidase (blue/white screening)
(Estimated time: 20 min)
Materials needed:
- 40 mg/ml X-gal in DMF
- SOC medium
- pre-warmed LB agar plate
- face mask
- The principle is that X-Gal (5-bromo-4-chloro-3-indolyl-b-D-galactopyranoside) turns blue when reacts with beta-galactosidase.
- Mix 20 ul X-Gal 20 mg/ml and 80 ul SOC and spread on LB+amp plates (X-Gal and DMF are toxic, use face-mask for your safety!!! X-Gal is light-sensitive, remember to keep it in the dark, when possible)
- Incubate with 200 ul bacteria
- Result: blue colonies express LacZ, while white colonies don't.
