Plasmid

Part:BBa_K4058030:Design

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


AlcR-pHSN6A01


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 11753
    Illegal XbaI site found at 1853
    Illegal XbaI site found at 2618
    Illegal XbaI site found at 6881
    Illegal SpeI site found at 13916
    Illegal PstI site found at 1678
    Illegal PstI site found at 2961
    Illegal PstI site found at 5952
    Illegal PstI site found at 7246
    Illegal PstI site found at 7792
    Illegal PstI site found at 9214
    Illegal PstI site found at 9418
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 11753
    Illegal NheI site found at 2651
    Illegal NheI site found at 3109
    Illegal NheI site found at 8458
    Illegal SpeI site found at 13916
    Illegal PstI site found at 1678
    Illegal PstI site found at 2961
    Illegal PstI site found at 5952
    Illegal PstI site found at 7246
    Illegal PstI site found at 7792
    Illegal PstI site found at 9214
    Illegal PstI site found at 9418
    Illegal NotI site found at 3424
    Illegal NotI site found at 6873
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 11753
    Illegal BglII site found at 2924
    Illegal BglII site found at 8048
    Illegal BglII site found at 8738
    Illegal BamHI site found at 1214
    Illegal BamHI site found at 6850
    Illegal BamHI site found at 8342
    Illegal BamHI site found at 10380
    Illegal BamHI site found at 11477
    Illegal XhoI site found at 1113
    Illegal XhoI site found at 3874
    Illegal XhoI site found at 6842
    Illegal XhoI site found at 7666
    Illegal XhoI site found at 8848
    Illegal XhoI site found at 10012
    Illegal XhoI site found at 12577
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 11753
    Illegal XbaI site found at 1853
    Illegal XbaI site found at 2618
    Illegal XbaI site found at 6881
    Illegal SpeI site found at 13916
    Illegal PstI site found at 1678
    Illegal PstI site found at 2961
    Illegal PstI site found at 5952
    Illegal PstI site found at 7246
    Illegal PstI site found at 7792
    Illegal PstI site found at 9214
    Illegal PstI site found at 9418
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 11753
    Illegal XbaI site found at 1853
    Illegal XbaI site found at 2618
    Illegal XbaI site found at 6881
    Illegal SpeI site found at 13916
    Illegal PstI site found at 1678
    Illegal PstI site found at 2961
    Illegal PstI site found at 5952
    Illegal PstI site found at 7246
    Illegal PstI site found at 7792
    Illegal PstI site found at 9214
    Illegal PstI site found at 9418
    Illegal NgoMIV site found at 1528
    Illegal NgoMIV site found at 5377
    Illegal NgoMIV site found at 8610
    Illegal NgoMIV site found at 14653
    Illegal AgeI site found at 12950
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 5666
    Illegal BsaI.rc site found at 4453
    Illegal SapI site found at 2764
    Illegal SapI site found at 7584
    Illegal SapI.rc site found at 1200
    Illegal SapI.rc site found at 9051


Design Notes

This construct contains the AlcR promoter from the Caddick et. al (1998) with the addition of a translational enhancer extracted from the sequence for pHSN6A01. This promoter is inserted before the dCas9 gene in pHSN6A01 to put dCas9 under ethanol inducible control.

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


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

Caddick, M., Greenland, A., Jepson, l. et al. An ethanol inducible gene switch for plants used to manipulate carbon metabolism. Nat Biotechnol 16, 177–180 (1998). https://doi-org.subzero.lib.uoguelph.ca/10.1038/nbt0298-177