DNA

Part:BBa_K4858002:Design

Designed by: Sriram Sai Garapati   Group: iGEM23_Michigan   (2023-10-02)


F2RL3 Quencher Probe (ProbeQ)


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:

Specificity: The utmost priority was granted to specificity. The probe was designed to ensure exclusive binding to the fluorescent probe in the absence of the SNP, mitigating potential off-target interactions.

Temperature Stability: To achieve consistency across fluorescence detection assays, the probe's melting temperature (Tm) was finely tuned to encompass typical assay temperature ranges.

TempG-F: -38.93 kcal/mol TempG-Sink: -58.6 kcal/mol F-Q: -76.96 kcal/mol Sink-Sink*: -23.97 kcal/mol

TempA-F: -78.56 kcal/mol TempA-Sink: -35.79 kcal/mol

deltaG = -RTlnQ


Source

Our probes are ordered from GenScript for experiments. We specified options that would allow us to attach the fluorophore, along with the associated sinks and quencher necessary for LAMP and PCR.

References

1. Hyman, L. B., Christopher, C. R., & Romero, P. A. (2022). Competitive SNP-lamp probes for rapid and robust single-nucleotide polymorphism detection. Cell Reports Methods, 2(7), 100242. https://doi.org/10.1016/j.crmeth.2022.100242