Based on a three fluorescent channel digital PCR method, the mutation sites of human EGFR gene can be precisely detected. A specific TAQMAN probe is used, aiming at loss (L858R, L861Q, and E19) of EGFR gene and T790M quadruple mutation area, 22 mutation sites related with EGFR-TKI drug sensitivity and drug resistance are simultaneously detected in a reaction liquid in a single tube, and moreover,the template type can be judged according to the copy number of mutant DNA. Aiming at the mutation (loss of No.19 exon) of human EGFR gene, a special probe is designed, specifically modified repression nucleotide and wild DNA are combined so as to ensure that detected fluorescent microspheres are all signals of mutant DNA; and then the template type is judged according to the copy number of mutantDNA. The provided kit and method use human blood to carry out liquid active detection; by detecting the situation of mutation of human EGFR gene, the problems that the conventional EGFR gene mutationdetection is tedious and the sensitivity is low are solved, and the stability, specificity, and sensitivity of human EGFR gene mutation detection are all largely improved.