Coding

Part:BBa_K4477003:Design

Designed by: Miranda Khoury   Group: iGEM22_Virginia   (2022-09-06)
Revision as of 08:15, 12 October 2022 by Mirandakhoury (Talk | contribs) (→‎Design Notes)


IK17 (anti-oxLDL) scFv antibody fragment


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 486
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

Amino acid sequence reverse transcribed and codon optimized for expression in E. coli B (the parent strain of SHuffle).

His tag and TEV cleavage sites were included downstream of the start codon but upstream of the antibody coding sequence. The His tag ensured the entire polypeptide could be purified out of a total protein solution using immobilized metal ion affinity chromatography (IMAC), and the TEV cleavage site ensured that TEV could be used to cleave off the His tag once it was no longer needed.

For detailed annotations, see Team Virginia 2022's Benchling folder here: https://benchling.com/mrkhoury/f_/YR4zZSUb-virginia-igem-2022-dna-constructs/

Source

Amino acid sequences for IK17 light and heavy chains acquired from NCBI: heavy: https://www.ncbi.nlm.nih.gov/protein/AAK93956.1 light: https://www.ncbi.nlm.nih.gov/protein/AAK93957.1

Literature and other relevant citations: https://www.jacc.org/doi/full/10.1016/j.jacc.2011.07.017 https://web.expasy.org/abcd/ABCD_AH862

References

1. https://www.jacc.org/doi/full/10.1016/j.jacc.2011.07.017
2. https://pubmed.ncbi.nlm.nih.gov/14644097/