Team:Newcastle/Measurement/Methods
Alternative Roots
Materials and Methods
Materials and Methods used regarding Measurement, Automation and Reproducibility
Bacterial Strains.
Transformations with, and expression of, iGEM test devices and controls were carried out using chemically competent Escherichia coli DH5α. Competency was conferred using the MgCl-CaCl2 method (Sambrook and Russell 2001). Briefly, a single colony of DH5α was incubated in Leuria Bertoni (LB) broth overnight at 37°C with shaking at 220rpm. Overnight culture was diluted 1:100, further incubated until an optical density (OD600nm) of 0.3 – 0.6 was reached and then placed on ice for 30 minutes. Cells were centrifuged at 4000g for 5 minutes at 4°C, resuspended in 0.1M MgCl2 and incubated on ice for 30 minutes. The suspension was centrifuged again as before, resuspended in 0.1M CaCl2 and placed on ice for 30 minutes. Cells were spun down again, resuspended in 0.1M CaCl2 with 15% glycerol and frozen at -80°C.