Team:Bielefeld-CeBiTec/Notebook/CFPS
Cell-free Protein Synthesis
04/22: Transformation of electrocompetent E. coli KRX with BBa_K1365020 (sfGFP (Bs))
04/23: preparing glycerol stock of KRX with BBa_K1365020
04/24-04/25: Mini-prep of BBa_K1365020
04/28: Transformation of electrocompetent E. coli KRX with BBa_I746909 (T7-promoter, RBS, sfGFP, double terminator). Before plating on LB(Cm), 50 µl of 200 g∙L-1 Rhamnose-solution is added to the plate to induce T7-Polymerase. Inoculation of 10 ml preculture of simple KRX from plate. Overnight culturing, shaking vigorously at 220 rpm.
04/29: colonies of KRX with BBa_I746909 look greenish! 2.2 ml Overnight cultures from KRX are used to inoculate 100 ml for start OD600=0.144. Measuring of growth curves for a first estimation of appropriate moment for cell harvest (see image below)
Growth curves (duplicate) from E. coli KRX at 37 °C in 100 ml 2xTY+P-medium. 1 l shaking flasks without baffles were used. Shaking at 200 rpm.