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Team:TzuChiU Formosa/Notebook
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'''2009.10.06''' OmpC promoter transformation. | '''2009.10.06''' OmpC promoter transformation. | ||
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Plasmid pSB1A3 biopart from 96-well plated was dissolved in 10 ul PCR water, 2 ul was used for plasmid transformation into DH5α competent cells. | Plasmid pSB1A3 biopart from 96-well plated was dissolved in 10 ul PCR water, 2 ul was used for plasmid transformation into DH5α competent cells. | ||
| - | '''2009.10.09''' | + | '''2009.10.08''' Positive clone selection and plasmid isolation. |
| + | |||
| + | 4:30 pm ~ 4:50 pm | ||
| + | Pick single bacterial colony that had been transform with OmpC promoter containing plasmid and inoculate with 3ml LB medium containing Ampicillin and grow overnight (16-20 hours) in a 37℃ shaker incubator. | ||
| + | |||
| + | |||
| + | |||
| + | '''2009.10.09''' Colony PCR to confirm OmpC promoter and plasmid isolation. | ||
| + | |||
| + | 2:00 pm ~ 5:00 pm | ||
| + | 10 ul overnight culture was used for colony PCR. Briefly 10 ul overnight culture was centrifuged at 13,000 rpm for 10 min, supernatant was removed, 500 ul autoclaved water was added fllowed by 100C incubation for 20 min and chilled on ice. 5 ul was used for colony PCR in a total 20 ul reaction with OmpC promoter specific primers. 5 ul of the PCR products was used for agarose gel electrophoresis to verify the positive clone. | ||
| + | |||
| + | |||
| + | '''2009.10.11''' Prepare LB plates | ||
| + | |||
| + | 8:30 am ~12:00 noon | ||
| + | 500 ml LB-agar was prepared and autoclaved. When LB-agar was ready (temperature cold enough to hold, about 60°C), 500 ul ampicillin (50 mg/ml) and 500 ul kanamycin (50 mg/ml) were added and 14 LB-agar plates were prepared. | ||
| + | |||
| + | 4:10 pm ~ 4:30 pm pSB1A3 preparation | ||
| + | Three single colonies containing pSB1A3 plasmid were inoculated in 5ml LB medium containing 7μl ampicillin (50 mg/ml) and grow overnight (16-20 hours) in a 37℃ shaker incubator. | ||
| + | |||
| + | |||
| + | '''2009.10.11''' Stock bacterial preparation and plating new plates for future used. | ||
| + | 4:30 pm ~ 4:50 pm | ||
| + | Two glycerol stock were prepared as follow : | ||
| + | 0.5 ml overnight culture was added into 0.5 ml 20% glycerol stock in a 2 ml eppendorf tube, mix well and kept at -80C. | ||
| - | + | 4:35pm~4:50pm | |
| + | Select two OmpC promoter transformed single colonies and inoculate 5ml LB contain 7μl Ampicillin and grow overnight (16-20 hours) in a 37℃ shaker incubator. | ||
Revision as of 12:08, 21 October 2009
Notebook
Meeting
2009.03 IGEM recuiting Poster
People from different department groups together after they see the poster.
2009.03.14 Experience sharing
NYMU-Taipei 2008 IGEM team came to TCU and shared Experience with TZU-CHI university.
2009.05.07 Firt meeting
Introduction of IGEM and advisors.
2009.05.15 2nd meeting
Groups formed and started brain storming.
2009.05.28 3rd meeting
Brain stormung and two project out of 12 were voted.
2009.07~2009.09 English course
English conversation course on Wednesdays, Thursdays.
2009.08.09 Japanese friends visited
Interactive friendship with University of Tokyo, and Chiba University, Japan.
2009.08.31 4th meeting
Progress report of the two groups.
2009.09.14 5th meeting
chose 1 team out of 2 .
2009.09.19 6th meeting
Reference searching.
2009.09.23 7th meeting
Reference searching and Determined the experimental designs.
2009.09.30 8th meeting with adviser
Reference searching and experimental designs report to advisors.
2009.10.2 9th meeting
Duties assigned. Started WIKI editing
2009.10.11 10th meeting
Progress report and upload project, protocals to wiki.
2009.10.15 11th meeting
Progress report.
2009.10.17 12th meeting
Upload results, discussion, notebook to wiki.
2009.10.20 13th meeting
Progress report and modification of wiki with advisors.
Experiments
2009.10.06 OmpC promoter transformation.
Plasmid pSB1A3 biopart from 96-well plated was dissolved in 10 ul PCR water, 2 ul was used for plasmid transformation into DH5α competent cells.
2009.10.08 Positive clone selection and plasmid isolation.
4:30 pm ~ 4:50 pm Pick single bacterial colony that had been transform with OmpC promoter containing plasmid and inoculate with 3ml LB medium containing Ampicillin and grow overnight (16-20 hours) in a 37℃ shaker incubator.
2009.10.09 Colony PCR to confirm OmpC promoter and plasmid isolation.
2:00 pm ~ 5:00 pm 10 ul overnight culture was used for colony PCR. Briefly 10 ul overnight culture was centrifuged at 13,000 rpm for 10 min, supernatant was removed, 500 ul autoclaved water was added fllowed by 100C incubation for 20 min and chilled on ice. 5 ul was used for colony PCR in a total 20 ul reaction with OmpC promoter specific primers. 5 ul of the PCR products was used for agarose gel electrophoresis to verify the positive clone.
2009.10.11 Prepare LB plates
8:30 am ~12:00 noon 500 ml LB-agar was prepared and autoclaved. When LB-agar was ready (temperature cold enough to hold, about 60°C), 500 ul ampicillin (50 mg/ml) and 500 ul kanamycin (50 mg/ml) were added and 14 LB-agar plates were prepared.
4:10 pm ~ 4:30 pm pSB1A3 preparation Three single colonies containing pSB1A3 plasmid were inoculated in 5ml LB medium containing 7μl ampicillin (50 mg/ml) and grow overnight (16-20 hours) in a 37℃ shaker incubator.
2009.10.11 Stock bacterial preparation and plating new plates for future used.
4:30 pm ~ 4:50 pm Two glycerol stock were prepared as follow : 0.5 ml overnight culture was added into 0.5 ml 20% glycerol stock in a 2 ml eppendorf tube, mix well and kept at -80C.
4:35pm~4:50pm Select two OmpC promoter transformed single colonies and inoculate 5ml LB contain 7μl Ampicillin and grow overnight (16-20 hours) in a 37℃ shaker incubator.
2009.10.12 T-A cloning.
2009.10.13 T-Atransformation.
2009.10.14 Cloning PCR.
2009.10.16 T-A cloning.
2009.10.20 Digestion, Gel extration
2009.10.21 Gel extration
.
