The invention relates to polypeptide capable of specifically targeting HER2 (human epidermal growth factor receptor 2) protein and an application of the polypeptide. The polypeptide is shown by the following general formula: X1X2X3X4X5LX6X7X8NPX9X10X11X12X13. The invention also relates to a nucleotide sequence for coding the polypeptide, and an expression vector for expressing the polypeptide, and host cells; and the invention also relates to the polypeptide, a bivalent or a multivalent formed by the polypeptide, and a pharmaceutical composition which is formed by the polypeptide serving as a targeting polypeptide, a preparation capable of killing cancer cells, and an imaging preparation, as well as applications of the polypeptide, the bivalent or the multivalent, and the pharmaceutical composition. The polypeptide provided by the invention can achieve a targeting effect on positive tumor cells of HER2, is strong in selectivity, can be prepared by adopting a chemical synthesis method, is high in purity, small in molecular weight and strong in specificity, and is free of immunogenicity, safe and reliable.