Plasmid

Part:BBa_K1391025

Designed by: Jing Wei "Raymond" Liu   Group: iGEM14_MIT   (2014-10-17)

High sensor: L7Ae_miR-125b

This part is our completed high sensor capable of detecting the presence of miR-125b.

Usage and Biology

miRNA-125b is one of the four miRNAs we have identified through literature searches to be upregulated in the neurons of Alzheimer's disease patients. When miR-125b is present in the system, it binds to target sites on the sensor. This silences expression of the repressor, L7ae. This signal then goes on to activate an output response on a separate plasmid.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 1085
    Illegal EcoRI site found at 2314
    Illegal PstI site found at 730
    Illegal PstI site found at 1011
    Illegal PstI site found at 1440
    Illegal PstI site found at 1945
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 1085
    Illegal EcoRI site found at 2314
    Illegal PstI site found at 730
    Illegal PstI site found at 1011
    Illegal PstI site found at 1440
    Illegal PstI site found at 1945
    Illegal NotI site found at 402
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 1085
    Illegal EcoRI site found at 2314
    Illegal BglII site found at 1694
    Illegal BglII site found at 3016
    Illegal XhoI site found at 394
    Illegal XhoI site found at 2093
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 1085
    Illegal EcoRI site found at 2314
    Illegal PstI site found at 730
    Illegal PstI site found at 1011
    Illegal PstI site found at 1440
    Illegal PstI site found at 1945
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 1085
    Illegal EcoRI site found at 2314
    Illegal PstI site found at 730
    Illegal PstI site found at 1011
    Illegal PstI site found at 1440
    Illegal PstI site found at 1945
    Illegal NgoMIV site found at 645
    Illegal NgoMIV site found at 926
    Illegal NgoMIV site found at 1828
    Illegal AgeI site found at 774
    Illegal AgeI site found at 1206
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 2769
    Illegal BsaI.rc site found at 2917


A



B

C

High Sensor miR-125b in the presence and absence of siRNA-125b in HEK293 cells. Using red as our transfection marker and green as our output, we expected that the sensor, when exposed to the inhibiting siRNA, would show an increase in GFP output. The cytometry results in figure (A) shows that there is difference between the 'on' and 'off' state. This conclusion is reinforced by the microscopy in (B) and (C). On the left the sensor is in a low miRNA environment, and on the right the sensor is in a high miRNA environment.

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