Difference between revisions of "Part:BBa K2332010:Design"

(→‎References)
(→‎Source)
Line 13: Line 13:
 
Intimin protein sequence obtained from UniProtKB: [http://www.uniprot.org/uniprot/P43261#sequences P43261], reverse translated and codon optimised for E. coli.
 
Intimin protein sequence obtained from UniProtKB: [http://www.uniprot.org/uniprot/P43261#sequences P43261], reverse translated and codon optimised for E. coli.
 
Intimin for the cell surface display of our tag was truncated according to Wentzel et al., 2001.
 
Intimin for the cell surface display of our tag was truncated according to Wentzel et al., 2001.
−
SpyTag was obtained from: [https://parts.igem.org/Part:BBa_K1159201 BBa_K1159201]
+
SpyTag was obtained from: [https://parts.igem.org/Part:BBa_K1159201 BBa_K1159201]. The DNA sequence was synthesised by Integrated DNA Technologies (IDT)
  
 
===References===
 
===References===

Revision as of 21:55, 22 October 2017


Intimin'-SpyTag


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 801
    Illegal NgoMIV site found at 1542
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

We decided to fuse SpyTag to the N-terminal of a truncated version of Intimin as this has been previously proven to function effectively for cell surface display of up to 30kDa passenger proteins (Wentzel et al., 2001). We also included a linker between Intimin and SpyTag to facilitate mobility of SpyTag on the cell surface.

Source

Intimin protein sequence obtained from UniProtKB: [http://www.uniprot.org/uniprot/P43261#sequences P43261], reverse translated and codon optimised for E. coli. Intimin for the cell surface display of our tag was truncated according to Wentzel et al., 2001. SpyTag was obtained from: BBa_K1159201. The DNA sequence was synthesised by Integrated DNA Technologies (IDT)

References

Zakeri B, Fierer J, Celik E, Chittock E, Schwarz-Linek U, Moy V et al. Peptide tag forming a rapid covalent bond to a protein, through engineering a bacterial adhesin. Proceedings of the National Academy of Sciences. 2012;109(12):E690-E697.

Wentzel A, Christmann A, Adams T, Kolmar H. Display of Passenger Proteins on the Surface of Escherichia coli K-12 by the Enterohemorrhagic E. coli Intimin EaeA. Journal of Bacteriology. 2001;183(24):7273-7284.