Colorectal cancer is a malignant tumor with high incidence and mortality worldwide. Chemotherapy serves as the core treatment for adjuvant therapy after surgery and advanced-stage patients; however, its application is often limited by drug resistance and dose-limiting toxicities (such as diarrhea, neurotoxicity, mucositis, etc.). In recent years, the gut microbiota, recognized as the “second genome” of the human body, has been confirmed to have a close bidirectional interaction with chemotherapeutic drugs: on the one hand, chemotherapeutic drugs can reshape the structure of the gut microbiota; on the other hand, the microbiota and their metabolites can significantly affect the efficacy and toxicity of chemotherapeutic drugs. This review aims to systematically elaborate on the core mechanisms underlying the bidirectional interaction between chemotherapy and gut microbiota, and focuses on exploring precise intervention strategies targeting the microbiota to reverse drug resistance and alleviate adverse effects. It holds great significance for achieving clinical translation and formulating personalized treatment plans.