Part:BBa_K2605003

A2UCOE

1.2kb A2UCOE fragment from the human HNRPA2B1-CBX3 (RNP/A2UCOE) locus

Usage and Biology

Eukaryotic transgene expression is highly influenced by the spread of heterochromatin, nearby enhancers, and promoter methylation, which in combination leads to highly variable levels of expression and even complete gene-silencing. To achieve stable and localization of integration-independent transgene expression, Ubiquitous Chromatin-Opening Elements (UCOEs), can be used (Neville et al. 2017, Romanova & Noll, 2017). UCOEs are methylation-free CpG islands found in the promoter regions of housekeeping genes in eukaryotic cells and when UCOEs are linked to a transgene promoter, they have been shown to increase expression levels as well as resist gene-silencing for hundreds of cell generations (Williams et al. 2005).

The 8kb UCOE from the human HNRPA2B1-CBX3 (RNP/A2UCOE) locus can be shortened down to a core 1.2kb fragment that functions as effectively as the whole fragment. The sequence for this 1.2kb fragment was taken from literature (Zhang et al., 2010).

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

References

Neville, J., Orlando, J., Mann, K., McCloskey, B., Antoniou, M. (2017). Ubiquitous Chromatin-opening Elements (UCOEs): Applications in biomanufacturing and gene therapy. Biotechnology Advances, 35(5), 557-564.

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

Williams, S., Mustoe, T., Mulcahy, T., Griffiths, M. Simpson, D., Antoniou, M., Irvine, A., Mountain, A., Crombie, R. (2005). CpG-island fragments from the HNRPA2B1/CBX3genomic locus reduce silencing and enhance transgene expression from the hCMV promoter/enhancer in mammalian cells. BMC Biotechnology, 5(17).

Zhang, F., Frost, A.R., Blundell, M.P., Bales, O., Antoniou, M.N., Thrasher, A.J. (2010). A ubiquitous chromatin opening element (UCOE) confers resistance to DNA methylation-mediated silencing of lentiviral vectors. Molecular Therapy, 18(9), 1640-49.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 73
  Illegal XhoI site found at 134
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]