The invention discloses a protein-based wood adhesive which is characterized by being made of the following raw materials in part by weight: 100 parts of water, 5 to 50 parts of protein-containing raw materials, 10 to 14 parts of polyamide epichiorobydrin (PAE), 5 to 15 parts of modifier, 1 to 15 parts of acidity regulator, 0 to 15 parts of stabilizer, 0 to 10 parts of humectant, 0 to 10 parts of anti-freeze, 0.1 to 10 parts of preservatives, 0 to 10 parts of mold inhibitor, 0 to 10 parts of enzyme inhibitor, and 0 to 10 parts of filling. The protein-containing raw materials contain over 10% of coarse protein, and the modifier is one of or a mixture of several of polyacrylamide, polyvinyl alcohol, polyvinyl acetate, borax, ammonium thiosulfate, carbamide and isocyanate. The invention also discloses a preparation method of the protein-based wood adhesive. The invention adopts common chemical materials and adopts a break-through normal-temperature material-mixing technique, which uses water as solvent. The adhesive has the advantages of high stickiness, evenness and convenience in adhesive application and stable product quality.