Imperial/Wet Lab/Lab Notebook/2007-08-14
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= 14 August 2007 = | = 14 August 2007 = | ||
+ | ==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== | ||
+ | #Picked a BL21(DE3) colony from Tet plate | ||
+ | #Innoculated in 100 ml of 2xYT + Tet medium | ||
+ | #Incubated overnight at 37 °C | ||
+ | |||
+ | ==Cloning of Biobricks== | ||
+ | #Placed 5 ml of LB in a 15 ml tube | ||
+ | #Added appropriate antibiotics into the tube | ||
+ | #Picked a colony from the fresh overnight plate | ||
+ | #Innoculated the colony in the LB media | ||
+ | #Incubated for 6 hours during the day | ||
+ | #*2x 6 Biobricks cloned | ||
+ | |||
+ | ==Cloning of Biobricks (for Midiprep)== | ||
+ | #Placed 50 ml of LB in a 250 ml conical flask | ||
+ | #Innoculated 1 ml of culture into LB media in flask | ||
+ | #Incubated overnight at 37 °C | ||
+ | |||
+ | <font size=-2> | ||
+ | *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] | ||
+ | </font> | ||
+ | |||
+ | ==Pilot Preparation of Vesicles== | ||
+ | |||
+ | *The suspension prepared the [[Imperial/Wet Lab/Lab Notebook/2007-08-13 | 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 100μl 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. | ||
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Latest revision as of 12:42, 26 October 2007
14 August 2007
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
- Picked a BL21(DE3) colony from Tet plate
- Innoculated in 100 ml of 2xYT + Tet medium
- Incubated overnight at 37 °C
Cloning of Biobricks
- Placed 5 ml of LB in a 15 ml tube
- Added appropriate antibiotics into the tube
- Picked a colony from the fresh overnight plate
- Innoculated the colony in the LB media
- Incubated for 6 hours during the day
- 2x 6 Biobricks cloned
Cloning of Biobricks (for Midiprep)
- Placed 50 ml of LB in a 250 ml conical flask
- Innoculated 1 ml of culture into LB media in flask
- 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 100μl 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.