Composite

Part:BBa_K3755041:Design

Designed by: Kaijun Wang   Group: iGEM21_ShanghaiTech_China   (2021-10-01)


CMV enhancer+CMV promoter+GCaMP6m+SV40 PolyA signal


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 542
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 683
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1524
    Illegal SapI.rc site found at 1430


Design Notes

We used PCR to obtain GCaMP6m from pAAV.Syn.Flex.GCaMP6m.WPRE.SV40. Then we inserted it behind CMV promoter using pEGFP as the backbone.

Source

It's a fusion protein which consists of a circularly permuted GFP (cpGFP), calmodulin (CaM) and a calmodulin-binding peptide(CBP). We get pAAV.Syn.Flex.GCaMP6m.WPRE.SV40(Part:BBa_K3755032) containing it from Wei Shen' lab, Shanghaitech university.

References

[1]Chen TW, Wardill TJ, Sun Y, Pulver SR, Renninger SL, Baohan A, Schreiter ER, Kerr RA, Orger MB, Jayaraman V, Looger LL, Svoboda K, Kim DS. Ultrasensitive fluorescent proteins for imaging neuronal activity. Nature. 2013 Jul 18;499(7458):295-300. doi: 10.1038/nature12354. PMID: 23868258; PMCID: PMC3777791.