Two Types of Mismatches Introduced Catalytic Hairpin Assembly Assay for Sensitive COVID-19 Detection
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Polymerase chain reaction (PCR) is the gold standard technique for Coronavirus (COVID-19) testing. However, PCR is time-consumed and labor-intensive due to enzymatic amplification. Accordingly, the catalytic hairpin assembly (CHA) has been proposed for enzyme-free signal-amplified nucleic acid detection. In this work, we designed a CHA assay with two hairpin DNAs that can be assembled by cascade strand displacement initialized by the N2-gene target. Two types of mismatch were introduced to increase the signal-to-noise. The first type of mismatch was introduced in the stem of the first hairpin DNA to promote strand displacement and increase the signal. The second mismatch was introduced in the final complex to reduce the background signal by suppressing the breathing effect. The detection limit was 5.9 times lowered with two types of mismatches. This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIT) (2020R1A2C2004168).