Team:HokkaidoU Japan/RBS/Methods

From 2013.igem.org

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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).

Revision as of 23:23, 28 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.