Primers were designed using Primer Premier 5.0, with two fusion sites with a high detection frequency targeted to generate an amplicon fragment size of 200 ~ 300 bp. The forward primers were designed based on HPV sequences, and the reverse primers were designed based on human genome sequences. B primers and C primers are displayed in Supplementary Table S11 .
Sequences were amplified using Phoenix™ Hot Start Taq DNA Polymerase (Enzymatics) in a PCR reaction mix containing 4 µL 5X Phoenix Hot Start Taq Reaction Buffer, 2 µL dNTPs (2.5 µM), 0.5 µL each of forward and reverse primer (10 µM), 0.2 µL Phoenix™ Hot Start Taq DNA Polymerase (500 U), 1 µL template DNA (10 ng), and 12.3 µL Nuclease-Free Water (not DEPC-treated), for a total volume of 20 µL. The PCR amplification conditions were as follows: 5 min at 95 °C; 35 cycles at 94 °C for 30 s, 60 °C for 60 s for annealing, and 72 °C for 60 s; and a final 72 °C for 1 min. PCR products were visualized via agarose gel electrophoresis and cleaned-up using AMPure XP beads (Beckman Coulter, Miami, FL, USA).
Sequences were amplified using Phoenix™ Hot Start Taq DNA Polymerase (Enzymatics) in a PCR reaction mix containing 4 µL 5X Phoenix Hot Start Taq Reaction Buffer, 2 µL dNTPs (2.5 µM), 0.5 µL each of forward and reverse primer (10 µM), 0.2 µL Phoenix™ Hot Start Taq DNA Polymerase (500 U), 1 µL template DNA (10 ng), and 12.3 µL Nuclease-Free Water (not DEPC-treated), for a total volume of 20 µL. The PCR amplification conditions were as follows: 5 min at 95 °C; 35 cycles at 94 °C for 30 s, 60 °C for 60 s for annealing, and 72 °C for 60 s; and a final 72 °C for 1 min. PCR products were visualized via agarose gel electrophoresis and cleaned-up using AMPure XP beads (Beckman Coulter, Miami, FL, USA).