Difference between revisions of "Part:BBa K649201:Experience"
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'''[Method]'''<br / > | '''[Method]'''<br / > | ||
1. Strain JM2.300 with both Pbad/araC-Cre and PlacIQ-lox2272-RFP-lox2272-GFP in it was cultured at 37℃ in 3ml of LB medium containing ampicillin (6 µL) and kanamycin(3.6 µL). Same kind of strain having only PlacIQ-lox2272-RFP-lox2272-GFP was cultured in same volume of LB containing kanamycin (3.6 µL) and carbenicillin(4 µL). These were cultured until OD 1.6. | 1. Strain JM2.300 with both Pbad/araC-Cre and PlacIQ-lox2272-RFP-lox2272-GFP in it was cultured at 37℃ in 3ml of LB medium containing ampicillin (6 µL) and kanamycin(3.6 µL). Same kind of strain having only PlacIQ-lox2272-RFP-lox2272-GFP was cultured in same volume of LB containing kanamycin (3.6 µL) and carbenicillin(4 µL). These were cultured until OD 1.6. | ||
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2. Each cultured medium was 6 times diluted in the medium and three samples (3 ml each) were dispensed from those. Among them, two were induced by arabinose (2 M, 75 µL). | 2. Each cultured medium was 6 times diluted in the medium and three samples (3 ml each) were dispensed from those. Among them, two were induced by arabinose (2 M, 75 µL). | ||
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3. After 30 min from induction, 1000 times dilution and 100000 times dilution of each sample were plated and incubated in 37°C about 12 hours. The florescence of plates was examined by FLC and it was taken picture. About the rest of the samples, strains were harvested by centrifugation and suspended by adding 1 mL of PBS (phosphate-buffered saline). The last OD of PBS solution was approximately 0.4. We dispensed 700 µL of each suspension into a disposable tube through a cell strainer, and fluorescence intensity of each cell was measured with a flow cytometer of Becton, Dickinson and Company. | 3. After 30 min from induction, 1000 times dilution and 100000 times dilution of each sample were plated and incubated in 37°C about 12 hours. The florescence of plates was examined by FLC and it was taken picture. About the rest of the samples, strains were harvested by centrifugation and suspended by adding 1 mL of PBS (phosphate-buffered saline). The last OD of PBS solution was approximately 0.4. We dispensed 700 µL of each suspension into a disposable tube through a cell strainer, and fluorescence intensity of each cell was measured with a flow cytometer of Becton, Dickinson and Company. | ||
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4. In the same way as 3, 9 more samples were examined at period 1hr, 2hr, and 4hr. | 4. In the same way as 3, 9 more samples were examined at period 1hr, 2hr, and 4hr. | ||
Revision as of 22:44, 3 October 2011
Applications of BBa_K649201
[Sample]
| sample | arabinose | |
|---|---|---|
| 1 | PlacIQ-lox2272-RFP-lox2272-GFP(pSB3K3) PBAD/araC-Cre(pSB1A2) |
+ |
| 2 | PlacIQ-lox2272-RFP-lox2272-GFP(pSB3K3) PBAD/araC-Cre(pSB1A2) |
- |
| 3 | PlacIQ-lox2272-RFP-lox2272-GFP(pSB3K3) : negative control |
+ |
[Method]
1. Strain JM2.300 with both Pbad/araC-Cre and PlacIQ-lox2272-RFP-lox2272-GFP in it was cultured at 37℃ in 3ml of LB medium containing ampicillin (6 µL) and kanamycin(3.6 µL). Same kind of strain having only PlacIQ-lox2272-RFP-lox2272-GFP was cultured in same volume of LB containing kanamycin (3.6 µL) and carbenicillin(4 µL). These were cultured until OD 1.6.
2. Each cultured medium was 6 times diluted in the medium and three samples (3 ml each) were dispensed from those. Among them, two were induced by arabinose (2 M, 75 µL).
3. After 30 min from induction, 1000 times dilution and 100000 times dilution of each sample were plated and incubated in 37°C about 12 hours. The florescence of plates was examined by FLC and it was taken picture. About the rest of the samples, strains were harvested by centrifugation and suspended by adding 1 mL of PBS (phosphate-buffered saline). The last OD of PBS solution was approximately 0.4. We dispensed 700 µL of each suspension into a disposable tube through a cell strainer, and fluorescence intensity of each cell was measured with a flow cytometer of Becton, Dickinson and Company.
4. In the same way as 3, 9 more samples were examined at period 1hr, 2hr, and 4hr.
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