SOE by PCR
SOE by PCR was carried out using a T100 Thermal Cycler (Bio-Rad, Hercules, CA, USA) for 20 cycles, each of 1 min at 98°C, 1 min at 55°C, and 2 min at 70°C. The reaction was carried out in a 50 µL volume containing 2.5 U Phusion High-Fidelity DNA polymerase (Pfu; Thermo Fisher Scientific), 200 µM dNTPs, 1 µL of primer mix (1.5 pmol per primer), and 5 µL of 10× Pfu buffer.