The invention relates to a tumor targeting polypeptide and a preparation method thereof. The amino acid sequential structure of the polypeptide of the invention is CFPVPGHDLVC. The preparation method comprises the following steps: co-culturing a pC89 phage peptide library for expressing 108 protein polypeptides with different sequences and human breast cancer cell strain MDA-MB-231 repeatedly for many times; screening phages which express specific polypeptides and can penetrate a cell membrane and enter cytoplasm and/or nucleus, amplifying the phages in vitro and carrying out DNA sequencing, and deducing exogenous amino acid sequences inserted on the phages; co-culturing the screened specific polypeptide phages, other human tumor cells and normal cells in vitro, and detecting the specificity of cell tumor of the polypeptide phages; and according to the polypeptide sequences and the detecting result of the cell specificity, artificially synthesizing the tumor targeting polypeptide. The invention has potential clinical application values for specific vectors of breast cancer targeting gene therapy, and also provides powerful technical support for screening polypeptides with affinity and specificity of other types of malignant tumor cell strains in the future. In addition, the polypeptide of the invention contains only 9-11 amino acid residues, can be synthesized easily, and has relatively small spatial position change, easy quality control of the polypeptide and convenient use.