Composite

Part:BBa_K200017:Design

Designed by: Royah Vaezi   Group: iGEM09_Imperial College London   (2009-10-15)
Revision as of 21:47, 19 October 2009 by Royah (Talk | contribs)

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RBS + OtsB


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 473
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

OtsB (BBa_K200006) was obtained from PCR from Escherichia Coli using BL21(DE3) as the genomic DNA template and the Pfu Ultra II enzyme. Primers were designed to include overhangs coding for XbaI and SpeI recognition sites in order to allow the gene to be BioBricked according to the BioBrick Standard. The primers are as follows:

Forward PCR primer containing XbaI overhang (bold) for BioBricking:
GCTCTAGATGACAGAACCGTTAACCG
Recomended Temperatures for PCR : 54.5C(without overhang) and 64.8C(with overhang)

Reverse PCR primer containing SpeI overhang (bold) for BioBricking:
GGACTAGTATTAGATACTACGACTAAACGAC
Recomended Temperatures for PCR : 54.7C(without overhang) and 64.2C(with overhang)

Biobrick Overhang shown in Bold


Source

PCR of OtsB from Escherichia Coli BL21(DE3), ligated to RBS extracted from Registry

References