DNA Extraction and OVA257-264 PCR Amplification

DNA was extracted from LLC-OVA cell lines utilizing the FastPure Cell/Tissue DNA Isolation Mini Kit (Vazyme, DC102-01). PCR amplifications were then conducted using specific primers, ensuring equivalent DNA quantities in each PCR reaction.
A 10 μl PCR mixture was prepared, comprising 1 μl of genomic DNA, 0.5 μl of each primer, 5 μl of 2 × Taq Master Mix (Vazyme, P112-01), and 3.5 μl of DNase/RNase-free ddH2O. The PCR parameters were set as follows: initial denaturation at 94 °C for 3 min, followed by 35 cycles of denaturation at 94 °C for 30 s, annealing at 60 °C for 35 s, and extension at 72 °C for 35 s. The reaction was concluded with a final extension at 72 °C for 5 min.
Following amplification, the PCR products were subjected to electrophoresis on a 1.0% agarose gel stained with Ultra GelRed (Vazyme, GR501-01) and visualized under ultraviolet light. The DL5000 DNA marker (Vazyme, MD102-01) was used to determine the molecular weight of the PCR products. The primers of OVA257-264^{102 (link)} were synthesized by Tsingke Biotechnology Co., Ltd (Beijing, China). The sequences of the primers are listed in Supplementary Table 2.