Plasmid

Part:BBa_K4058024:Design

Designed by: Sarah Cumberland   Group: iGEM21_Guelph   (2021-10-20)


ICE2 sgRNA A in pHSN6A01


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 7833
    Illegal XbaI site found at 2961
    Illegal SpeI site found at 9996
    Illegal PstI site found at 2032
    Illegal PstI site found at 3326
    Illegal PstI site found at 3872
    Illegal PstI site found at 5294
    Illegal PstI site found at 5498
    Illegal PstI site found at 6740
    Illegal PstI site found at 12904
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 7833
    Illegal NheI site found at 4538
    Illegal SpeI site found at 9996
    Illegal PstI site found at 2032
    Illegal PstI site found at 3326
    Illegal PstI site found at 3872
    Illegal PstI site found at 5294
    Illegal PstI site found at 5498
    Illegal PstI site found at 6740
    Illegal PstI site found at 12904
    Illegal NotI site found at 2953
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 7833
    Illegal BglII site found at 4128
    Illegal BglII site found at 4818
    Illegal BamHI site found at 2930
    Illegal BamHI site found at 4422
    Illegal BamHI site found at 6460
    Illegal BamHI site found at 7557
    Illegal XhoI site found at 2922
    Illegal XhoI site found at 3746
    Illegal XhoI site found at 4928
    Illegal XhoI site found at 6092
    Illegal XhoI site found at 8657
    Illegal XhoI site found at 9751
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 7833
    Illegal XbaI site found at 2961
    Illegal SpeI site found at 9996
    Illegal PstI site found at 2032
    Illegal PstI site found at 3326
    Illegal PstI site found at 3872
    Illegal PstI site found at 5294
    Illegal PstI site found at 5498
    Illegal PstI site found at 6740
    Illegal PstI site found at 12904
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 7833
    Illegal XbaI site found at 2961
    Illegal SpeI site found at 9996
    Illegal PstI site found at 2032
    Illegal PstI site found at 3326
    Illegal PstI site found at 3872
    Illegal PstI site found at 5294
    Illegal PstI site found at 5498
    Illegal PstI site found at 6740
    Illegal PstI site found at 12904
    Illegal NgoMIV site found at 1457
    Illegal NgoMIV site found at 4690
    Illegal NgoMIV site found at 10733
    Illegal AgeI site found at 9030
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1746
    Illegal BsaI.rc site found at 533
    Illegal SapI site found at 3664
    Illegal SapI.rc site found at 5131

Design Notes

pHSN6A01 was a gift from Qi-Jun Chen (Addgene plasmid # 50586 ; http://n2t.net/addgene:50586 ; RRID:Addgene_50586)

The gRNA sequence was designed using the guide RNA design software CHOPCHOP. The parameters were set to find optimal CRISPR/Cas9 target sites for CRISPR activation. Target sequences were selected based on the predicted binding efficiency provided by CHOPCHOP.

Source

pHSN6A01 was a gift from Qi-Jun Chen (Addgene plasmid # 50586 ; http://n2t.net/addgene:50586 ; RRID:Addgene_50586)


References

A CRISPR/Cas9 toolkit for multiplex genome editing in plants. Xing HL, Dong L, Wang ZP, Zhang HY, Han CY, Liu B, Wang XC, Chen QJ. BMC Plant Biol. 2014 Nov 29;14(1):327. 10.1186/s12870-014-0327-y PubMed 25432517

Labun, K., Montague, T. G., Krause, M., Torres Cleuren, Y. N., Tjeldnes, H., & Valen, E. CHOPCHOP v3: expanding the CRISPR web toolbox beyond genome editing. Nucleic Acids Research (2019).