CRISPR-Cas12a-Mediated Growth of Gold Nanoparticles for DNA Detection in Agarose Gel

The rapid, simple, and sensitive detection of nucleic acid biomarkers plays a significant role in clinical diagnosis. Herein, we develop a label-free and point-of-care approach for isothermal DNA detection through the trans-cleavage activity of CRISPR-Cas12 and the growth of gold nanomaterials in agarose gel. The presence of the target can activate CRISPR-Cas12a to cleave single-stranded DNA, thus modulating the length and number of DNA sequences that mediate the growth of gold nanoparticles (AuNPs) or gold nanorods (AuNRs). Due to the extraordinary plasmonic property of gold nanomaterials, they present characteristic absorption/color after the growth with unique shapes. The sensing strategy is applied to detect BRCA-1, a biomarker related to breast cancer, with limits of detection of 1.72 pM (AuNP-based) and 2.07 pM (AuNR-based). AuNPs/AuNRs can be immobilized in agarose gels that display different colors in the presence of target DNA sequences. The agarose gel-based test allows for a readout by the naked eye or the RGB value with a smartphone. The approach is isothermal and label-free without any surface modification of nanomaterials, which holds great potential for the detection of nucleic acids in clinical applications.