Fig. 7
Workflow diagram for detection. The process begins with DNA extraction using the QIAGEN extraction kit, which requires 20 min. Next, DNA amplification is performed, either through RPA (20 min) or PCR (90 min). Traditional PCR requires gel preparation and, consequently, the samples must undergo electrophoresis. Additionally, a visualization system is necessary to observe the DNA within the gel. In some cases, sequencing may be required for further verification. Otherwise, RPA amplification proteins and Cas protein incubation occur at 37 °C, eliminating the need for temperature variation. Following amplification, a CRISPR/LbCas12a cleavage step (10 min) is performed, in which the guide RNA and Cas protein identify and cleave the target region (cis cleavage). Finally, a fluorescence signal is generated during trans cleavage by the Cas protein, taking 20 min to produce visible fluorescence on the TropD-Detector. Figure created BioRender.com
