The invention relates to a method for constructing a sinocyclocheilus rhinocerous heart cell line. The method comprises the following steps: 1, obtaining of sinocyclocheilus rhinocerous heart tissues, wherein sinocyclocheilus rhinocerous is sterilized through combination of penicillin and ethyl alcohol; 2, primary culture, wherein a culture solution M199 which contains glutamine, penicillin, streptomycin, streptomycin sulphate and amphotericin is adopted for culture, and the osmotic pressure of the culture solution is 280-330 mOsm / l; 3, secondary culture, wherein when the tenth generation is reached, cell culture fluid is replaced with base culture fluid, and the sinocyclocheilus rhinocerous heart cell line is constructed successfully. According to the method for constructing the sinocyclocheilus rhinocerous heart cell line, the obtained sinocyclocheilus rhinocerous heart cell line is in the fibrous form and can conduct continuous passage and be directly applied to research of biological characteristics, requirements of genetic resource preservation and theoretical research conducted on the rare species, namely the sinocyclocheilus rhinocerous, can be satisfied, meanwhile, the heart cell line is the first heart cell line of cave fishes in China, and the foundation is laid for cave adaptability research at the cellular level.