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TALEA-dife

Part:BBa_K1189021:Design

Designed by: Chris Wintersinger, Denny Hoang, Taylor Remondini   Group: iGEM13_Calgary   (2013-09-17)
Revision as of 17:42, 1 November 2013 by Cmwinter (Talk | contribs)

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TALE-A ferritin di-subunit direct fusion w/ hisTag under lacI promoter, and linker


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 2654
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 3656

Design notes

This part was created by fusing the heavy chain (BBa_K1189025, P02794 [UniParc]) and light chains (BBa_K1189024, P02792 [UniParc]) of human ferritin together with TALE-A (BBa_K1189022). It is expressed under the LacI promoter (BBa_J04500) and has a his-tag for protein purification. BBa_K1189021 is expressed under a strong RBS and the LacI promoter.

Source

We synthesized heavy ferrtin (BBa_K1189019), light ferritin (BBa_K1189020), and a codon optimized TALE-A (BBa_K1189022) from a commerical synthesis company. These parts were optimized for expression in E. coli K12. The ferritin di-subunit (BBa_K1189018) and TALE-A (BBa_K1189022) were extracted via PCR, the pSB1C3 backbone was isolated with PCR, and these three linear fragments were assembled into BBa_K1189021 using isothermal assembly.

References

  • Chasteen, N. D., & Harrison, P. M. (1999). Mineralization in ferritin: an efficient means of iron storage. Journal of structural biology, 126(3), 182-194.
  • Dehal, P. K., Livingston, C. F., Dunn, C. G., Buick, R., Luxton, R., & Pritchard, D. J. (2010). Magnetizable antibody‐like proteins. Biotechnology journal, 5(6), 596-604.