Coding

Part:BBa_K2020052

Designed by: Andrea Hoeltken, Carolina Bonerath, Vroni Czotscher   Group: iGEM16_Aachen   (2016-10-15)
Revision as of 12:43, 17 October 2016 by Andrea hoeltken (Talk | contribs) (→‎Full Set of DMNBS synthtetases)


DMNBS-Synthetase for use in E.coli

This is a DMNBS-synthetase to be used as a orthogonal synthetase in E.coli. This part can be used together with the cognate tRNA BBa_K2020042 to incorporate DMNBS in response to an amber stop codon.

Compared to incorporation of tyrosine with wild type Mj Y-Synthetase in response to an amber codon:

  • Incorporation Efficiency: % (DMNBS incorporation value)
  • Incorporation Fidelity: %(descrimination againt tyrosine)

Full Set of DMNBS synthtetases

Usage and Biology

incorporation of ncAA

Assembly in a synthetase plasmid for incorporation of ncAA

alt text

Most synthetases are used with low copy plasmids (e.g. pACYC). Assemble the tRNA and the synthetase into a low copy plasmid, each one with an own promoter and one terminator for both. (See picture). If your application is not for incorporation into a protein but for use with a second plasmid, make shure to use replicons from different incompatibility groups, eg. ColE1 and p15A and different selection markers. A second plasmid could be the flourescent reporter plasmid pFRY for the purpose of determining fidelity and efficiacy of synthetases for ncAA.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 84
    Illegal SapI.rc site found at 877


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Categories
Parameters
None