Difference between revisions of "Part:BBa K649201:Experience"

 
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This experience page is provided so that any user may enter their experience using this part.<BR>Please enter
 
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how you used this part and how it worked out.
 
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===Applications of BBa_K649201===
 
===Applications of BBa_K649201===
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We prepared a competent cell JM2.300 into which Pbad/araC-Cre(pSB1A2, BBa_I718008) had been constructed. Subsequently, the BioBrick was constructed into the cell. The strain was grown in a 3ml liquid culture, and 75μl of 2M arabinose was added to induce Cre expression. For the control experiments we used the same strain without arabinose induction and a JM2.300 strain which was induced with arabinose and had only our BioBrick.  All the strains were cultured each for periods of 0.5, 1, 2, and 4 hours, and in each case the florescence levels were measured by flow cytometer and FLC.
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table
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We measured the fluorescence levels to prove that recombination only occurs in presence of Cre recombinase. On the sample with the Pbad/araC-Cre construction, we found that recombination occurred when arabinose was added. We could also observe recombination occurred when arabinose was not added, which can be explained due to a leaking in the Pbad/araC promoter. Opposite to this result, when we measured the levels of the sample without the Pbad/araC-Cre construction, we found that the GFP levels were far lower than those of the sample with the Pbad/araC-Cre construction. This clearly proves that our lox constructions, both in BBa_K649201 and BBa_K649202, respond correctly to the effects of Cre recombinase.
  
 
===User Reviews===
 
===User Reviews===

Revision as of 13:08, 3 October 2011

Applications of BBa_K649201

We prepared a competent cell JM2.300 into which Pbad/araC-Cre(pSB1A2, BBa_I718008) had been constructed. Subsequently, the BioBrick was constructed into the cell. The strain was grown in a 3ml liquid culture, and 75μl of 2M arabinose was added to induce Cre expression. For the control experiments we used the same strain without arabinose induction and a JM2.300 strain which was induced with arabinose and had only our BioBrick. All the strains were cultured each for periods of 0.5, 1, 2, and 4 hours, and in each case the florescence levels were measured by flow cytometer and FLC. 

table

We measured the fluorescence levels to prove that recombination only occurs in presence of Cre recombinase. On the sample with the Pbad/araC-Cre construction, we found that recombination occurred when arabinose was added. We could also observe recombination occurred when arabinose was not added, which can be explained due to a leaking in the Pbad/araC promoter. Opposite to this result, when we measured the levels of the sample without the Pbad/araC-Cre construction, we found that the GFP levels were far lower than those of the sample with the Pbad/araC-Cre construction. This clearly proves that our lox constructions, both in BBa_K649201 and BBa_K649202, respond correctly to the effects of Cre recombinase.

User Reviews

UNIQ345d8fedd7d1bc74-partinfo-00000000-QINU UNIQ345d8fedd7d1bc74-partinfo-00000001-QINU