The present invention provides a method and a reagent for enrichment of circulating tumor DNA, the method comprising the steps of mixing a water phase and an oil phase and shaking the mixture to prepare an emulsion PCR reaction system, and performing emulsion PCR amplification, wherein the water phase comprises peripheral blood plasma DNA as template DNA, a forward primer and a reverse primer, dNTPs, a PCR buffer and a DNA polymerase, the peripheral blood plasma DNA having adapter sequences connected to both ends thereof, and the forward primer and the reverse primer being complementary to the adapter sequences at the two ends respectively; separating the water phase from the oil phase following the emulsion PCR amplification to obtain a PCR amplification product in the water phase; and capturing circulating tumor DNA in the PCR amplification product in the water phase by using a probe sequence that specifically binds to the circulating tumor DNA. The method of the present invention is capable of performing single-molecule high-fidelity ultramicro parallel amplification and effectively capturing peripheral blood plasma ctDNA to provide adequate amount of ctDNA to be used for subsequent sequencing detection.
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