PM_2.5 induces oxidative/antioxidant system imbalance and excessive release of reactive oxygen species (ROS) and produces toxic effects and irreversible damage to the genetic material including chromosomes and DNA. Biochanin A (BCA), an isoflavone with strong antioxidant activity, effectively intervenes against PM_2.5-induced oxidative damage. The X-ray repair cross-complementary protein 1 (XRCC1)/BER pathway involves DNA damage repair caused by oxidative stress. This paper aims to explore the mechanism of BCA alleviating oxidative DNA damage caused by PM_2.5 by establishing the in vitro cell model based on CRISPR/Cas9 technology and combining it with mechanism pathway research. The results showed that PM_2.5 exposure inhibited the expression of BER and NER pathway proteins and induced the overexpression of ERCC1. BCA showed an effective intervention in the toxicity of PM_2.5 in normal cells, rather than XRCC1 knock-out cells. This laid a foundation for further exploring the key role of XRCC1 in PM_2.5-caused oxidative damage and the BER/DNA damage repair pathway.