Part:BBa_K2506001:Design

IL-17RA_SynNotch_Gal4-VP64

Assembly Compatibility:

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

  Illegal NotI site found at 2416
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 273
  Illegal BglII site found at 796
  Illegal BglII site found at 2202
  Illegal BamHI site found at 618
  Illegal BamHI site found at 968
  Illegal BamHI site found at 2614
  Illegal XhoI site found at 1
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 2409
  Illegal BsaI.rc site found at 1861
  Illegal SapI site found at 1618
  

Design Notes

Starting from the N-terminus of the fusion protein, we use the extracellular domain of the IL17RA gene. Then, refering to the paper (Roybal KT, Williams JZ, et al. Cell 2016. doi: 10.1016/j.cell.2016.09.011.), we add Notch 1 gene sequence and replace the mouse domain with human domain. In the end, we combined GAl4-VP64 from Addgene with Notch 1 sequence.

Source

We get the DNA sequences of the IL-17RA and Notch 1 protein in Unipro. And we also find the DNA sequence of GAl4-VP64 from Addgene. IL-17RA http://www.uniprot.org/uniprot/Q96F46 Notch1 http://www.uniprot.org/uniprot/P46531 GAl4-VP64 http://www.addgene.org/85421/sequences/

References

1. Roybal KT, Williams JZ, et al. Engineering T Cells with Customized Therapeutic Response Programs Using Synthetic Notch Receptors. Cell 2016. doi: 10.1016/j.cell.2016.09.011.

2. Roybal KT, Rupp LJ. et al. Precision Tumor Recognition by T Cells With Combinatorial Antigen-Sensing Circuits. Cell. 2016. doi: 10.1016/j.cell.2016.01.011.

3. Kononenko AV, Lee NC. et al. Generation of a conditionally self-eliminating HAC gene delivery vector through incorporation of a tTAVP64 expression cassette. Nucleic Acids Res. 2015. doi: 10.1093/nar/gkv124.

4. Müller K, Zurbriggen MD, Weber W. An optogenetic upgrade for the Tet-OFF system. Biotechnol Bioeng. 2015. doi: 10.1002/bit.25562.

5. Chen Z, Barbi J, et al. The ubiquitin ligase Stub1 negatively modulates regulatory T cell suppressive activity by promoting degradation of the transcription factor Foxp3. Immunity. 2013. doi: 10.1016/j.immuni.2013.08.006.

6. Van Loosdregt J, Fleskens V, et al. Stabilization of the transcription factor Foxp3 by the deubiquitinase USP7 increases Treg-cell-suppressive capacity. Immunity. 2013. doi: 10.1016/j.immuni.2013.05.018.

7. Wang L, Kumar S, et al. Ubiquitin-specific Protease-7 Inhibition Impairs Tip60-dependent Foxp3+ T-regulatory Cell Function and Promotes Antitumor Immunity. EBioMedicine. 2016. doi: 10.1016/j.ebiom.2016.10.018.

8. Ren J, Li B. The Functional Stability of FOXP3 and RORγt in Treg and Th17 and Their Therapeutic Applications. Adv Protein Chem Struct Biol. 2017;107:155-189. doi: 10.1016/ bs.apcsb.2016.10.002. Epub 2016 Dec 15.

9. Chen X, Oppenheim JJ. Th17 cells and Tregs: unlikely allies. J Leukoc Biol. 2014 May;95(5):723-731. Epub 2014 Feb 21.