Team:HokkaidoU Japan/RBS/Methods

From 2013.igem.org

(Difference between revisions)
 
(6 intermediate revisions not shown)
Line 3: Line 3:
   <div id="common-header-bottom-background">
   <div id="common-header-bottom-background">
     <div class="wrapper">
     <div class="wrapper">
-
       <h1 id="common-header-title">Maestro E.coli</h1>
+
       <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>
       <img id="common-header-img" src="https://static.igem.org/mediawiki/2013/e/ea/HokkaidoU2013_Maestro_Header.png">
       <img id="common-header-img" src="https://static.igem.org/mediawiki/2013/e/ea/HokkaidoU2013_Maestro_Header.png">
Line 17: Line 17:
<p>
<p>
   We constructed new RBS family, SD2, SD4, SD6, SD8.
   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).
+
   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).
Line 24: Line 24:
<div class="fig fig800">
<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">
-
   <div>fig.1: oligos; RED: enhancer sequence, BLUE: SD sequence.</div>
+
   <div><span class="bold">fig.1: oligos.</span> RED: enhancer sequence, BLUE: SD sequence.</div>
</div>
</div>
<div class="fig fig400 para">
<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">
-
   <div>fig.2: RBS construction</div>
+
   <div><span class="bold">fig.2: RBS construction.</span></div>
</div>
</div>
<div class="fig fig400 para">
<div class="fig fig400 para">
-
   <img src="https://static.igem.org/mediawiki/2013/2/28/HokkaidoU_RBS_methods3_400.png">
+
   <img src="https://static.igem.org/mediawiki/2013/1/1a/HokkaidoU2013_RBS_methods3revision_400.png">
-
   <div>fig.3: our parts</div>
+
   <div><span class="bold">fig.3: our parts.</span></div>
</div>
</div>
<div class="clearfix"></div>
<div class="clearfix"></div>

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.