Difference between revisions of "Part:BBa K649200:Experience"
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===Applications of BBa_K649200=== | ===Applications of BBa_K649200=== | ||
| − | In in vitro assay, the part was made linear using restriction enzymes. Cre recombinase was added to the linear DNA and incubated for 0.5, 2, and 4 hours. Images of the experiments have been added below.<br /><br /> | + | In ''in vitro'' assay, the part was made linear using restriction enzymes. Cre recombinase was added to the linear DNA and incubated for 0.5, 2, and 4 hours. Images of the experiments have been added below.<br /><br /> |
'''[Sample]'''<br /> | '''[Sample]'''<br /> | ||
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</tr> | </tr> | ||
<tr> | <tr> | ||
| − | <td>PlacIQ-lox2272- | + | <td>PlacIQ-''lox2272-gfp-lox2272''(pSB1C3)</td> |
<td>50 ng/μl × 4 μl</td> | <td>50 ng/μl × 4 μl</td> | ||
</tr> | </tr> | ||
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'''[Method]''' | '''[Method]''' | ||
<br /> | <br /> | ||
| − | 1. The part on pSB1C3 was made linear by EcoRV restriction site which is far from lox sites. <br /><br /> | + | 1. The part on pSB1C3 was made linear by EcoRV restriction site which is far from ''lox'' sites. <br /><br /> |
| − | 2. Three identical samples and one negative control | + | 2. Three identical samples and one negative control supplied with ddH2O instead of Cre-recombinase had been prepared and those were incubated in 37°C for 0.5hr, 2hr and 4hr respectively. Negative control was incubated for 4hr.<br /><br /> |
| − | 3. After a period of time, the samples were kept in -20°C until the whole samples are arranged. The result was checked by electrophoresis(0.8% EtBr(+) agarose gell, 100V, 400mA, 30min) | + | 3. After a period of time, the samples were kept in -20°C until the whole samples are arranged. The result was checked by electrophoresis<br />(0.8% EtBr(+) agarose gell, 100V, 400mA, 30min). |
| − | + | <br /> | |
| + | <br /> | ||
| + | [[Image:in vitro_lox_gfp_lox.png|thumb|center|500px|<br/>|''in vitro'' assay of K649200<br />negative control, 0.5hr, 2hr, and 4hr from left]] | ||
| + | <br /> | ||
| + | These image of the electrophoresis experiment shows that there are several bands in samples to which Cre was added, which indicates that excision of the ''lox'' sites successfully occurred. | ||
===User Reviews=== | ===User Reviews=== | ||
Latest revision as of 05:11, 29 October 2011
Applications of BBa_K649200
In in vitro assay, the part was made linear using restriction enzymes. Cre recombinase was added to the linear DNA and incubated for 0.5, 2, and 4 hours. Images of the experiments have been added below.
[Sample]
| sample | volume |
|---|---|
| PlacIQ-lox2272-gfp-lox2272(pSB1C3) | 50 ng/μl × 4 μl |
| 10× Cre recombination Buffer | 1 μl |
| ddH2O | 4 μl |
| Cre recombinase | 1,000 units/ml × 1 μl |
| total | 10 μl |
[Method]
1. The part on pSB1C3 was made linear by EcoRV restriction site which is far from lox sites.
2. Three identical samples and one negative control supplied with ddH2O instead of Cre-recombinase had been prepared and those were incubated in 37°C for 0.5hr, 2hr and 4hr respectively. Negative control was incubated for 4hr.
3. After a period of time, the samples were kept in -20°C until the whole samples are arranged. The result was checked by electrophoresis
(0.8% EtBr(+) agarose gell, 100V, 400mA, 30min).
These image of the electrophoresis experiment shows that there are several bands in samples to which Cre was added, which indicates that excision of the lox sites successfully occurred.
User Reviews
UNIQ157c821b623c8dbd-partinfo-00000000-QINU UNIQ157c821b623c8dbd-partinfo-00000001-QINU