Composite

Part:BBa_K3407022:Design

Designed by: Javier Navarro Delgado   Group: iGEM20_TUDelft   (2020-10-23)


Short hairpin RNA (shRNA) potential trigger of RNAi. Transcription controlled under T7 promoter.


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

This biobrick represents the DNA template for the transcription of a 65 nt shRNA, including the promoter. Trancription was performed with T7 RiboMAX kit. The dsRNA sequence was taken from pUC57-OriLR-deGFP plasmid (BBa_K3407006). It corresponds to 27nt from the eGFP gene (nt 78 to 105), and is the one recognised and processed by Dicer to form siRNA. The shRNA has a single-stranded RNA (ssRNA) containing Fox-1 RBD (BBa_K3407004) RNA target sequence “UUGCAUGUU” [1] in its loop. The dsRNA region contains a GG overhang that would represent the overhang left when mini-3 (BBa_K3407002) cleaves a tshRNA (See iGEM TU Delft 2020, Design - Future perspectives), and that represents a good substrate to process the shRNA by Dicer into a siRNA.


References

Ordered List

  1. Auweter, S., Fasan, R., Reymond, L., Underwood, J., Black, D., Pitsch, S. and Allain, F., 2020. Molecular Basis Of RNA Recognition By The Human Alternative Splicing Factor Fox-1.