Composite

Part:BBa_K2812005:Design

Designed by: Guido Oerlemans, Maxime van den Oetelaar and Mariska Brüls   Group: iGEM18_TU-Eindhoven   (2018-10-03)


Coding sequence for trunctated Lysostaphin with HlyA and His6-tag regulated by T7-promoter


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 1389
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 164
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

The sequence for the T7 promotor was has been taken from BBa_K525998. The initial starting sequence for truncated lysostaphin was taken from BBa_K748002. The amino acid sequence for HlyA was taken from uniprot (UniProtKB - Q1R2T5 (Q1R2T5_ECOUT)), residue 807-1024.1 This is the C-terminal signal sequence, the N-terminal catalytic domain responsible for lysis of red blood cells is not included. The thrombin substrate sequence was obtained from Waugh, 20112 and all three components were linked via two flexible GGGGS linkers in between. This amino acid sequence was used to generate a codon optimized sequence for expression in E. coli.

Source

The designed sequence for lysostaphin-thrombin linker-HlyA-His-tag was synthesized de novo by IDT.

References

https://www.uniprot.org/uniprot/Q1R2T5