Regulatory

Part:BBa_K2605004

Designed by: Kaitlin Schaaf   Group: iGEM18_Calgary   (2018-10-09)


Chicken beta-globin insulator

1.2kb insulator from Gallus gallus (chicken) beta-globin locus.

Usage and Biology

Insulators are DNAse‐I hypersensitive CpG‐rich structure that block unrelated enhancer-promoter interactions and when lying between a promoter and enhancer pair, the insulator prevents the interactions between these elements (Pikaart, M.J., Recillas-Targa, F., & Felsenfeld, G, 1998). A large problem in mammalian eukaryotic biology is that promoters and enhancers contained within integrated cassettes could possibly interact with endogenous promoters or enhancers. The use of insulators can help reduce the amount of interactions by creating an isolated pocket for transgene expression (Romanova & Noll, 2017). When flanking a transgene, insulators also function as a road-block to the spread of heterochromatin, which helps maintain stably active expression of that gene (Pikaart, M.J., Recillas-Targa, F., & Felsenfeld, G, 1998).

The most well-characterized insulator is a 1.2kb element from the 5' end of the Gallus gallus (chicken) beta-globin locus and it functions to block enhancer function and to prevent heterochromatin spread (Pikaart, M.J., Recillas-Targa, F., & Felsenfeld, G, 1998). The insulator sequence was found on NCBI (GenBank: U78775.2)

This part contains a 5' BmtI restriction site and 3' BamHI restriction site so it can be used within our Multiple-Cloning Site flanked by FRT sites Part BBa_K2605002 system.

References

Pikaart, M.J., Recillas-Targa, F., Felsenfeld, G. (1998). Loss of transcriptional activity of a transgene is accompanied by DNA methylation and histone deacetylation and is prevented by insulators. Genes and Development, 12, 2852-2862.

Romanova, N., Noll, T. (2017). Engineered and Natural Promoters and Chromatin‐Modifying Elements for Recombinant Protein Expression in CHO Cells. Biotechnology Journal, 13(3). Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 11
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 637
    Illegal BamHI site found at 1249
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 152
  • 1000
    COMPATIBLE WITH RFC[1000]


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