Difference between revisions of "Part:BBa K2332052:Design"

 
(→‎Design Notes)
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===Design Notes===
 
===Design Notes===
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Removal of ATG from GFP for a more efficient expression of this fusion protein
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We decided to use 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 included a linker between Intimin' and GFP and between GFP and SpyTag to facilitate mobility of GFP and SpyTag on the cell surface. The stop codon in GFP was removed for the generation of this fusion protein.
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===Source===
 
===Source===

Revision as of 23:38, 22 October 2017


Intimin'-GFP-SpyTag (constitutive)


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal SpeI site found at 2826
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
    Illegal SpeI site found at 2826
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal SpeI site found at 2826
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal SpeI site found at 2826
    Illegal NgoMIV site found at 862
    Illegal NgoMIV site found at 1603
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 2697


Design Notes

We decided to use 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 included a linker between Intimin' and GFP and between GFP and SpyTag to facilitate mobility of GFP and SpyTag on the cell surface. The stop codon in GFP was removed for the generation of this fusion protein.

Source

Intimin protein sequence obtained from: http://www.uniprot.org/uniprot/P43261#sequences And reverse translated using: http://www.bioinformatics.org/sms2/rev_trans.html Intimin for the cell surface display of our tag was truncated according to Wentzel et al., 2001 SpyTag: BBa_K1159201 GFPmut 3b: Part:BBa_E0040

References