Difference between revisions of "Part:BBa K1632011"
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− | <span style="margin-left: 10px;">In order to assay the function of our FimE, we added a GFP coding sequence on the downstream of the <i>fim</i> switch.The <i>fim</i> switch[default ON](wild-type)_gfp (BBa_K1632007) emitts fluorescence when expressed, while the <i>fim</i> switch [default OFF](wild-type)_gfp(BBa_K1632008) does not emit florescence when expressed. We also added PBAD/''araC'' on the upstream of fimE.PBAD/''araC''_fimE (BBa_K1632013) can induce the expression of FimE in the presence of arabinose. We co-transformed a <i>fim</i> | + | <span style="margin-left: 10px;">In order to assay the function of our FimE, we added a GFP coding sequence on the downstream of the <i>fim</i> switch.The <i>fim</i> switch[default ON](wild-type)_gfp (BBa_K1632007) emitts fluorescence when expressed, while the <i>fim</i> switch [default OFF](wild-type)_gfp(BBa_K1632008) does not emit florescence when expressed. We also added PBAD/''araC'' on the upstream of fimE.PBAD/''araC''_fimE (BBa_K1632013) can induce the expression of FimE in the presence of arabinose. We co-transformed a <i>fim</i> switch_gfp and a PBAD/''araC''_fim recombinase in the E. coli DH5alpha strain. We measured the fluorescence intensity of the cells induced by different concentraions of arabinose.<br> |
[[Image:Tokyo_Tech_FimE_assay_Results.png |thumb|center|700px|<b>Fig. 5. </b>Histogram of the samples measured by flow cytometer]]<br> | [[Image:Tokyo_Tech_FimE_assay_Results.png |thumb|center|700px|<b>Fig. 5. </b>Histogram of the samples measured by flow cytometer]]<br> |
Revision as of 10:07, 18 September 2015
fimE (wild-type)
The fim switch is inverted by FimE.The FimE protein inverts the fim switch predominantly in the ON-to-OFF direction.
In order to assay the function of our FimE, we added a GFP coding sequence on the downstream of the fim switch.The fim switch[default ON](wild-type)_gfp (BBa_K1632007) emitts fluorescence when expressed, while the fim switch [default OFF](wild-type)_gfp(BBa_K1632008) does not emit florescence when expressed. We also added PBAD/araC on the upstream of fimE.PBAD/araC_fimE (BBa_K1632013) can induce the expression of FimE in the presence of arabinose. We co-transformed a fim switch_gfp and a PBAD/araC_fim recombinase in the E. coli DH5alpha strain. We measured the fluorescence intensity of the cells induced by different concentraions of arabinose.
From the experimental results,our fimE inverted the fim switch[default ON](wild-type) from ON-to-OFF but did not invert the fim switch[default OFF](wild-type) from the OFF state, depending on the concentration of arabinose.
For more information, see [http://2015.igem.org/Team:Tokyo_Tech/Project our work in Tokyo_Tech 2015 wiki].
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 22
- 1000COMPATIBLE WITH RFC[1000]