Difference between revisions of "Part:BBa K2332054:Design"

 
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===Design Notes===
 
===Design Notes===
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To obtain the fusion protein of GFP-SpyTag, we had to remove the stop codon of GFP. Parts were already codon optimized for E. Coli.
 
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We removed the stop codon in GFP and placed the SpyCatcher sequence after a linker sequence and included a HisTag for protein purification.
  
 
===Source===
 
===Source===
  
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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 SpyCatcher: BBa_K1159200 GFPmut 3b: Part:BBa_E0040
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SpyCatcher sequence was obtained from [https://parts.igem.org/Part:BBa_K1159200 BBa_K1159200] and GFP from:  
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[https://parts.igem.org/Part:BBa_E0040 BBa_E0040]. The DNA sequence was synthesised by Integrated DNA Technologies (IDT)
  
 
===References===
 
===References===
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1. 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.

Latest revision as of 23:47, 22 October 2017


GFP-SpyCatcher (constitutive)


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 723


Design Notes

We removed the stop codon in GFP and placed the SpyCatcher sequence after a linker sequence and included a HisTag for protein purification.

Source

SpyCatcher sequence was obtained from BBa_K1159200 and GFP from: BBa_E0040. The DNA sequence was synthesised by Integrated DNA Technologies (IDT)

References

1. 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.