Colormetric Detection of miRNA with DNA-Functionalized Gold Nanoparticles

Abstract

Early detection of cervical cancer is important for improving patient prognosis. The use of miRNAs are promising biomarkers for cancer screening because they are found in peripheral blood and their dysregulation is associated with cancer progression. However, rates of cervical cancer incidence and mortality are highest among low- and middle-income communities and current miRNA detection assays, like RT-PCR, require expensive equipment. To remedy this, our team is focusing on increasing accessibility of cancer screening by developing a cost-effectice assay that can be heated and quantified using a portable sensor. The presence of specific miRNA initiates ligation, rolling circle transcription, and the activation of Cas12a. Our assay employs a rolling circle transcription reaction, which activates a Cas12a enzyme when the miRNA target is present. Activated Cas12a cleaves the DNA linker conjugated to the gold nanoparticles (AuNPs). In the absence of Cas12a activation, the linker sequence promotes AuNP aggregation while the absence of the linker prevents aggregation. However, the Cas12a activation assay must be combined with the AuNP complex to ensure proper aggregation when linker is intact, and that there is a lack of aggregation when Cas12a cleaves the linker. The cleavage produces a colorimetric output which can then be quantified using a 3D printed portable detector.