Difference between revisions of "Part:BBa K3725040:Design"
 
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<strong>Design</strong>
 
<strong>Design</strong>
  
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The construction of a disease-specific biosensor required us to find a gene unique to the pathogen. When the switch turns on and GFP is expressed, we can confirm that the specific pathogen is present. For the detection of Phytophthora cryptogea, Lambert iGEM focused on the X24 gene. This gene was selected because it was required for pathogenicity and was unique to the species of interest. Biosafety note, the trigger sequence is not the full transcript sequence and therefore poses limited biosafety. We obtained the sequence via UniProt, an online database of protein sequences. Lambert iGEM used the code from Takahashi et. al provided by Megan McSweeney from the Styczynski Lab at the Georgia Institute of Technology to design the switch and trigger sequences on NUPACK. The team selected the pair from NUPACK with the lowest normalized ensemble defect (NED) to maximize the chances of successful compatibility. Once we obtained the sequences for the toehold pair, we constructed the toehold and trigger via SnapGene.
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The construction of a disease-specific biosensor required us to find a gene unique to the pathogen. When the switch turns on and GFP is expressed, we can confirm that the specific pathogen is present. For the detection of <i>Phytophthora cryptogea</i>, Lambert iGEM focused on the X24 gene. This gene was selected because it was required for pathogenicity and was unique to the species of interest. Biosafety note, the trigger sequence is not the full transcript sequence and therefore poses limited biosafety. We obtained the sequence via UniProt, an online database of protein sequences. Lambert iGEM used the code from Takahashi et. al provided by Megan McSweeney from the Styczynski Lab at the Georgia Institute of Technology to design the switch and trigger sequences on NUPACK. The team selected the pair from NUPACK with the lowest normalized ensemble defect (NED) to maximize the chances of successful compatibility. Once we obtained the sequences for the toehold pair, we constructed the toehold and trigger via SnapGene.
 
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<center>[[File:T--Lambert GA--T7PhytoTriggerCircularConstruct.png|500px]]</center>
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<center>[[File:T--Lambert GA-PhytoTriggerConstructDiagram.png|500px]]</center>
 
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<center><I>Figure 1: This graph compares the mean lux values of the C. elegans toehold switch, trigger, and the dual plasmid. The toehold switch and trigger have very similar lux values to LB and plain cells, whereas the dual plasmid has a higher lux value, indicating fluorescence.</I></center>
 
<center><I>Figure 1: This graph compares the mean lux values of the C. elegans toehold switch, trigger, and the dual plasmid. The toehold switch and trigger have very similar lux values to LB and plain cells, whereas the dual plasmid has a higher lux value, indicating fluorescence.</I></center>

Latest revision as of 21:35, 21 October 2021


T7 Fusarium Trigger

Design

The construction of a disease-specific biosensor required us to find a gene unique to the pathogen. When the switch turns on and GFP is expressed, we can confirm that the specific pathogen is present. For the detection of Phytophthora cryptogea, Lambert iGEM focused on the X24 gene. This gene was selected because it was required for pathogenicity and was unique to the species of interest. Biosafety note, the trigger sequence is not the full transcript sequence and therefore poses limited biosafety. We obtained the sequence via UniProt, an online database of protein sequences. Lambert iGEM used the code from Takahashi et. al provided by Megan McSweeney from the Styczynski Lab at the Georgia Institute of Technology to design the switch and trigger sequences on NUPACK. The team selected the pair from NUPACK with the lowest normalized ensemble defect (NED) to maximize the chances of successful compatibility. Once we obtained the sequences for the toehold pair, we constructed the toehold and trigger via SnapGene.

T--Lambert GA-PhytoTriggerConstructDiagram.png


Figure 1: This graph compares the mean lux values of the C. elegans toehold switch, trigger, and the dual plasmid. The toehold switch and trigger have very similar lux values to LB and plain cells, whereas the dual plasmid has a higher lux value, indicating fluorescence.


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

Design Considerations


Source

Part Source

References