Plasmid
TNY2AKUB

Part:BBa_K076000

Designed by: iGEM2008 Princeton team   Group: iGEM08_Princeton   (2008-10-29)

pLV-TRE-Ngn1-linker-EYFP-2A-mKate-Ubc-Bla

A plasmid allowing Dox-induced expression of the cell-fate regulator Ngn1 fused to an enhanced yellow fluorescent protein reporter, and a red fluorescent protein reporter tethered by a 2A linker. This construct not only allows us to see neuronal differentiation, but also allows for the verification of the 2A linker, with the result of yellow-labeled Ngn1 remaining in the nucleus and red-labeled mKate being exported to the cytoplasm. This result follows because since Ngn1 acts in the nucleus, and EYFP moves into the nucleus along with Ngn1 because the two are fused. The mKate protein remains in the cytoplasm because it is severed from the Ngn1-EYFP construct at the 2A sequence.


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 328
    Illegal EcoRI site found at 1452
    Illegal EcoRI site found at 3304
    Illegal XbaI site found at 2875
    Illegal XbaI site found at 3298
    Illegal SpeI site found at 2802
    Illegal SpeI site found at 8621
    Illegal SpeI site found at 9295
    Illegal PstI site found at 3921
    Illegal PstI site found at 11870
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 328
    Illegal EcoRI site found at 1452
    Illegal EcoRI site found at 3304
    Illegal NheI site found at 1572
    Illegal SpeI site found at 2802
    Illegal SpeI site found at 8621
    Illegal SpeI site found at 9295
    Illegal PstI site found at 3921
    Illegal PstI site found at 11870
    Illegal NotI site found at 865
    Illegal NotI site found at 9331
    Illegal NotI site found at 10053
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 328
    Illegal EcoRI site found at 1452
    Illegal EcoRI site found at 3304
    Illegal BglII site found at 626
    Illegal BglII site found at 4085
    Illegal BglII site found at 4151
    Illegal BglII site found at 4192
    Illegal BglII site found at 8384
    Illegal BglII site found at 9383
    Illegal BamHI site found at 1201
    Illegal XhoI site found at 2863
    Illegal XhoI site found at 9338
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 328
    Illegal EcoRI site found at 1452
    Illegal EcoRI site found at 3304
    Illegal XbaI site found at 2875
    Illegal XbaI site found at 3298
    Illegal SpeI site found at 2802
    Illegal SpeI site found at 8621
    Illegal SpeI site found at 9295
    Illegal PstI site found at 3921
    Illegal PstI site found at 11870
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 328
    Illegal EcoRI site found at 1452
    Illegal EcoRI site found at 3304
    Illegal XbaI site found at 2875
    Illegal XbaI site found at 3298
    Illegal SpeI site found at 2802
    Illegal SpeI site found at 8621
    Illegal SpeI site found at 9295
    Illegal PstI site found at 3921
    Illegal PstI site found at 11870
    Illegal NgoMIV site found at 843
    Illegal NgoMIV site found at 1095
    Illegal NgoMIV site found at 4780
    Illegal NgoMIV site found at 5813
    Illegal NgoMIV site found at 5874
    Illegal NgoMIV site found at 5988
    Illegal AgeI site found at 11654
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 4174
    Illegal BsaI site found at 9365
    Illegal BsaI.rc site found at 2114
    Illegal BsaI.rc site found at 3934
    Illegal BsaI.rc site found at 7516
    Illegal BsaI.rc site found at 9254
    Illegal BsaI.rc site found at 10881
    Illegal BsaI.rc site found at 13096
    Illegal SapI site found at 6433
    Illegal SapI.rc site found at 10017
    Illegal SapI.rc site found at 11067
    Illegal SapI.rc site found at 12478


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