Coding

Part:BBa_K2019004:Design

Designed by: John Lazar   Group: iGEM16_Northwestern   (2016-10-12)


Periplasm Directed saCas9 via DsbA Signaling Sequence (DsbA-saCas9)


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 783
    Illegal BglII site found at 936
    Illegal BglII site found at 1082
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 356
    Illegal AgeI site found at 2912
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

DsbA signaling sequence was ordered as a gblock with homology to the BBa_K2019000-BBa_R0040 expression system. BBa_K2019000-BBa_R0040 was linearized at the 5' end of saCas9. A 2 piece gibson assembly was then performed with the gblock and Cas9 expression backbone.


Source

DsbA sequence is from the supplementary information of the paper "The DsbA Signal Sequence Directs Efficient, Cotranslational Export of Passenger Proteins to the Escherichia coli Periplasm via the Signal Recognition Particle Pathway".

References