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Team:Groningen/26 July 2010
From 2010.igem.org
Hydrophobofilm
pushing coatings into a greener future
Week 30
Jorrit
Tested transformation of week 29 on SDS-page with silver staining on the CFE, medium & pellet samples, no chaplains visible.
Transformation B sub.
Inoculated NZ8900 ΔTasA in 10 ml MM, overnight @ 37°C shaker.
1 ml overnight culture NZ8900 ΔTasA in 10 ml fresh MM for 3 hours @ 37°C shaker.
Added approx. 1 µg plasmid E and H in two 2 ml tubes and incubated 30 min @ 37°C shaker.
Diluted in pre warmed TY (300 µl) and put for 45 min @ 37°C shaker.
Plated on LB-agar with antibiotic Cm5/Spec100 and grown @ 37°C (colonies @ 30/7).
Inoculated 4 H colonies and 4 E colonies in 5 ml TY with antibiotics Km5 ,Cm5 and Spec100 and put overnight @ 37°C shaker.
Two -80°C stocks were made of B. sub NZ8900 ΔTasA with pNZ8901_E and B. sub NZ8900 ΔTasA with pNZ8901_H.
Inoculated 4 Greiner tubes with pNZ9801 E6, E2, H1, H3 in 5 ml TY with antibiotic Km5 and Cm5.
10 µl of OD cultures (pNZ9801 E6, E2, H1, H3) was taken and grown , overnight @ 37°C shaker, in 5 ml TY with antibiotic Cm5/Spec100 and Cm5.
Done growth curve with OD- measurement.
Of all measurementperiodes were taken 2 ml samples, spinned down and put supernatant and cell pellet in freezer.
