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Team:BCCS-Bristol/Notebook/Week 4
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* Fiu still not being ligated on ANY backbone. | * Fiu still not being ligated on ANY backbone. | ||
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| + | [[Image:BCCS_Fhua_osme_midipreps.png|left|200px|frame|Standard 1kb NEB Ladder. Lanes 1-2 FhuA Midipreps, Lanes3-4 OsmE midipreps.]] | ||
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| + | [[Image:BCCS_Fhua_gfp_pcr.png|center|200px|frame|Standard 1kb NEB Ladder. Lanes 1-2 FhuA PCR fragment for Quick and Dirty assembly Method, Lanes3-4 GFP PCR fragments for quick and dirty assembly method.]] | ||
Revision as of 11:38, 9 October 2009
BCCS-Bristol
iGEM 2009
iGEM 2009
Week 4
- One more try to get Fiu protein on pSB1A3 backbone. NovaBlue transformed with ligation products of pSB1A3 and Fiu.
- Grew colonies for FhuA and OsmE midi preps.
- Carried out midi preps for FhuA and OsmE ( [FhuA]=378.9 ng/ul, [OsmE]=113.2 ng/ul).
- Transformed Nova Blue with biobrick part BBa_B0014 (double terminator)
- Culture colonies of BBa_B0014 transformants to get ready for mini prep.
- PCR FhuA and GFP for assembling together a quick and dirty fusion to be ready prior to BioScaffold arrival.
- Carried out colony growth (agar & liquid) for the AraC promoter (2 parts BBa_R0080 (lacking O2 region) and BBa_R0081 (the O2 region).
- Miniprepped DNA from cultures of BBa_R0080 and R0081.
- Attempted a 2-way ligation and of the BBa_R0080 and R0081 parts. The final product should be a 2-component biobrick on a plasmid in 5'-R0081-R0080-3' direction which can be transformed into cells.
- R0081 part was purified by gel extraction after being treated by double digest. R0080 and R0081 were ligated together and used to transform NovaBlue cells.
- First attempt to make the AraC promoter from the two parts failed.
- Fiu still not being ligated on ANY backbone.
Standard 1kb NEB Ladder. Lanes 1-2 FhuA Midipreps, Lanes3-4 OsmE midipreps.
Standard 1kb NEB Ladder. Lanes 1-2 FhuA PCR fragment for Quick and Dirty assembly Method, Lanes3-4 GFP PCR fragments for quick and dirty assembly method.
