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Team:Chiba/Calendar-Home/21 August 2008
From 2008.igem.org
20 August 2008 <|> 22 August 2008
Contents |
Laboratory work
Team:Input
(-->20/8)
Inoculated transformants for 12 hours in LB 2mL containing Ampicillin.
- BBa_R0051(2007)
- BBa_R0051(2006)
- BBa_J06650(2007)
- BBa_J06650(2006)
- BBa_J22136(2007)
- BBa_J22141(2007)
BBa_J22141:After inoculated for 11 hours, added 20μl IPTG and cultured one hour.
- UV irradiation test (plate phase)
- two plasmids from(JW1908)
- (Repressor)-Ptrc-LuxR-Plux-cI-colE1-Amp-
- (Reporter)-PcI-GFP-p15a-Cm-
- Inoculated (Reporter) culture from glycerol stock(-80°C freezer) in 2mL LB-Ampicillin, Chloramphenicol medium and LB-Ampicillin, Chloramphenicol, 100nmAHL mediumfor 12h, at 37°C.
- We diluted 105-fold, and plated 20ul of the resulting solution to (LB-Amp,Cm)(LB-Amp,cm,AHL100nM) plates each other.
- We cultured for 12h at 37°C.
- We iraddiated UV each plate. Wavelength:254nm,distance from the UV lamp to the cultures were 14cm.
- We performed the same operations on plates following 9 or 21h of UV
exposure.
- Colonies from both plates were picked and cultured in 2mL of
LB-Amp,Cm or LB-Amp, Cm, 100nMAHL for 12h at 37°C.
- We diluted 105-fold, and plated 20ul of the
resulting solution to (LB-Amp,Cm)(LB-Amp,cm,AHL100nM) plates each other.
- Colony count
| no AHL | AHL100nM | |
| 9h | 606 | 453 |
| 12h | 146 | 151 |
result
GFP fluorescence was observed from plates without AHL.
Team:Communication
| DNA Template | 1 |
| dNTP mix(μL) | 10 |
| Foward Primer(μL) | 5 |
| Reverse Primer(μL) | 5 |
| DNA polymerase TAQ(μL) | 1 |
| Thermopol Buffer(μL) | 5 |
| dH2O(μL) | 28 |
| TOTAL(μL) | 50 |
- 95°C,5min -> ( 95°C,1min -> 52°C,1min -> 72°C,1min )･･･25cycles -> 72°C,10min -> 6°C
- --->Gel Check
|
- Transformation
- competent cells : XL10G
- BBa_S03154(2007)
- BBa_S03154(2006)
- BBa_I9026(2007)
- BBa_I9026(2006)
- BBa_I9030(2007)
- BBa_I9030(2006)
--->(23/8)Digestion
Team:Output
- BBa_J32007(2007)-->no colony-->no colony(2 times)-->colony(3times)
- BBa_B0034(2007)-->colony
