Team:HokkaidoU Japan/RBS/Methods

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       <h1 id="common-header-title">Maestro E.coli</h1>
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       <h1 id="common-header-title">Maestro <span class="italic">E. coli</span></h1>
       <h2 id="common-header-subtitle">RBS</h2>
       <h2 id="common-header-subtitle">RBS</h2>
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   We constructed new RBS family, SD2, SD4, SD6, SD8.
   We constructed new RBS family, SD2, SD4, SD6, SD8.
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   These RBSs have Enhancer sequence (GCTCTTTAACAATTTATCA) and SD sequence (SD2:GG, SD4:GAGG, SD6:AGGAGG, SD8:TAAGGAGG).
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   These RBSs have enhancer sequence (GCTCTTTAACAATTTATCA) and SD sequence (SD2:GG, SD4:GAGG, SD6:AGGAGG, SD8:TAAGGAGG).
   We constructed SD8 from synthetic oligos (forward:SD8-f, reverse:SD8-r).
   We constructed SD8 from synthetic oligos (forward:SD8-f, reverse:SD8-r).
   We constructed SD2, SD4, SD6 by PCR (forward:EX-f, reverse:SD2-r, SD4-r, SD6-r, template:SD8).
   We constructed SD2, SD4, SD6 by PCR (forward:EX-f, reverse:SD2-r, SD4-r, SD6-r, template:SD8).
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<div class="fig fig800">
   <img src="https://static.igem.org/mediawiki/2013/a/ab/HokkaidoU_RBS_methods1_800.png">
   <img src="https://static.igem.org/mediawiki/2013/a/ab/HokkaidoU_RBS_methods1_800.png">
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   <div>Fig.1: oligos; RED: enhancer sequence, BLUE: SD sequence.</div>
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   <div><span class="bold">fig.1: oligos.</span> RED: enhancer sequence, BLUE: SD sequence.</div>
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<div class="fig fig400 para">
   <img src="https://static.igem.org/mediawiki/2013/9/9e/HokkaidoU_RBS_methods2_400.png">
   <img src="https://static.igem.org/mediawiki/2013/9/9e/HokkaidoU_RBS_methods2_400.png">
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   <div>Fig.2: RBS construction</div>
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   <div><span class="bold">fig.2: RBS construction.</span></div>
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   <img src="https://static.igem.org/mediawiki/2013/2/28/HokkaidoU_RBS_methods3_400.png">
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   <img src="https://static.igem.org/mediawiki/2013/1/1a/HokkaidoU2013_RBS_methods3revision_400.png">
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   <div>Fig.3: our parts</div>
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   <div><span class="bold">fig.3: our parts.</span></div>
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<h2>Assay</h2>
<h2>Assay</h2>
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   We ligated TetR repressible promoter (pTET), each of the new RBSs', LacZ&alpha; and double terminator.
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   We ligated TetR repressible promoter (pTet), each of the new RBSs', LacZ&alpha; and double terminator.
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   Using this construct we performed β-Galactosidase assay.
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   Using this construct we performed &beta;-Galactosidase assay.
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<a href="https://2013.igem.org/Team:HokkaidoU_Japan/RBS"><div class="arrow-div"></div><span>RBS Top</span></a>
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<a href="https://2013.igem.org/Team:HokkaidoU_Japan/RBS/Results"><div class="arrow-div"></div><span>Results</span></a>
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Latest revision as of 02:52, 29 October 2013

Maestro E. coli

RBS

RBS family parts

We constructed new RBS family, SD2, SD4, SD6, SD8. These RBSs have enhancer sequence (GCTCTTTAACAATTTATCA) and SD sequence (SD2:GG, SD4:GAGG, SD6:AGGAGG, SD8:TAAGGAGG). We constructed SD8 from synthetic oligos (forward:SD8-f, reverse:SD8-r). We constructed SD2, SD4, SD6 by PCR (forward:EX-f, reverse:SD2-r, SD4-r, SD6-r, template:SD8).

fig.1: oligos. RED: enhancer sequence, BLUE: SD sequence.
fig.2: RBS construction.
fig.3: our parts.

Assay

We ligated TetR repressible promoter (pTet), each of the new RBSs', LacZα and double terminator. Using this construct we performed β-Galactosidase assay.