UF-PCR, which is based on the same principle as that of SYBR Green qPCR, was used by replacing SYBR Green with EvaGreen as an intercalating dye (Shin et al., 2022 (link)). PCR analysis was performed using a GENECHECKER UF-150 Ultra-fast Real-Time PCR system (Genesystem, Daejeon, Republic of Korea) with Rapi:chip. The final reaction volume was 10 µL consisting of 5 µL SSO Fast EvaGreen Supermixes (Bio-Rad Laboratories, Berkeley, CA, USA), 1 µL each of forward and reverse primers (0.8 µM), and 2 µL DNA template (50 ng). All PCR reactions were performed under the following conditions: one cycle of initial denaturation at 95 °C for 1 min, 40 cycles of denaturation at 95 °C for 5 s, annealing at 54 °C for 5 s, extension at 72 °C for 5 s, and one cycle of elongation at 72 °C for 5 s. No template control (NTC) was used as a negative control. The amplicons were analyzed using a microchip electrophoresis system, MCE-202 MultiNA (Shimazu, Nishinokyo, Kyoto, Japan) with MultiNA software v1.12 (Shimazu).