2x5 mL LB medium were inoculated with 50 µL DH5Alpha, aliquoted for a preculture and incubated overnight (16,5h) at 37°C.

250ml were inoculated with 5ml of preculture and incubated until they reached an OD of 0,5. After centrifugation, the cell pellet was washed with HTP buffer and resuspended in 3ml HTP buffer. In the end, 225µl DMSO was added to an end concenctration of 6-7%. The 50µl aliquots were frozen in N2(l) and stored at -80°C.

1µl plasmid with a amp-resistence was put on ice for 10min. A heat shock was induced for 45s at 42°C. The sample was put on ice for another 10min and then incubated in 200µl LB for 1h before it was plated onto LB-, ampicilin- and canamycin-plates.

50ml LB-medium (with ampicilin) were inoculated with E.coli BL21 PlyS and grown overnight.

Plates from 26.03.2014

Inoculation of LB-plates with E.Coli DH5Alpha with PSG1164.

Miniprep according to the miniprep kit protocol (Omega) with 2x6ml preculture.

DNA-concentration:

• 1. 233,2 ng/µl
• 2. 261,3 ng/µl

Digestion scheme:

Incubate samples for 60 min at 37°C, then induce a heat shock for 2 min at 45°C. 5µl marker and 10µl of the samples are loaded onto the SDS-gel.