2.5 Evaluation of the sensitivity and fluorescence intensity of CRISPR Cas12a Detection
CRISPR Cas12a detection assays contained 50nM LbCas12a, 100 nM crRNA, 0.5 μl RNase inhibitor, 2 μl NEBuffer 2.1, FAM-BHQ1-labeled ssDNA reporter and various amount of dsDNA template p3xFLAG-CMV-7.1-p72. The sample without dsDNA plasmid template was set as negative control. Reactions were incubated at a constant temperature of 37oC for 30 min and the fluorescent intensity was measured by ABI QuantStudio 5. In addition, fluorescence was observed with a UV transilluminator.