IGEM:IMPERIAL/2007/Notebook/2007-8-14

name=iGEM:IMPERIAL/2007/Notebook date=2007/09/15 view=threemonths format=%name/%year-%month-%day weekstart=7

Preparation of Stock Solutions

 * 1x 1 litre of 2xYT medium in Duran bottle
 * 3x 1 litre of 2xYT medium in conical flask
 * Sent to autoclave
 * 1x 10 ml of pre-incubation solution for preparation of cell extract
 * 6x 20 ml of 1mM IPTG stock

Preparation of Cell Extract

 * 1) Picked a BL21(DE3) colony from Tet plate
 * 2) Innoculated in 100 ml of 2xYT + Tet medium
 * 3) Incubated overnight at 37 °C

Cloning of Biobricks

 * 1) Placed 5 ml of LB in a 15 ml tube
 * 2) Added appropriate antibiotics into the tube
 * 3) Picked a colony from the fresh overnight plate
 * 4) Innoculated the colony in the LB media
 * 5) Incubated for 6 hours during the day
 * 6) *2x 6 Biobricks cloned

Cloning of Biobricks (for Midiprep)

 * 1) Placed 50 ml of LB in a 250 ml conical flask
 * 2) Innoculated 1 ml of culture into LB media in flask
 * 3) Incubated overnight at 37 °C


 * 2. BBa_I13422 [ptet-GFP]
 * 5. BBa_R0040 [ptet]
 * 7. BBa_F2620 [plux]
 * 8. BBa_F1610 [luxI]
 * 11. BBa_E7104 [pT7]
 * 13. BBa_B0015 [stop]
 * 14. BBa_R0051 [pcI]
 * 16. BBa_E0240 [GFP]
 * 17. BBa_I13507 [RFP]
 * 18. BBa_T9002 [plux-GFP]

Pilot Preparation of Vesicles

 * The suspension prepared the day before was used to produce vesicles enclosing Tris and NaCl buffer only.


 * There was a mistake in following the protocol: instead of preparing the interface with 2ml of the DOPC-mineral oil suspension and then adding 100&mu;l of emulsion to it, the emulsion was prepared according to protocol, and then 2ml of emulsion was used to create the interface.


 * Results: A sample was briefly looked at under a light microscope, but no vesicles were observed.