Colorimetric biosensors have attracted wide attention due to their low cost, simple operation, rapid response and good reproducibility. However, insufficient sensitivity limits their applications. This report describes the design of a colorimetric biosensor based on a three-step multiple signal amplification strategy to detect breast cancer-associated BRCA1 mutation. The capture unit, signal unit, and target DNA form a sandwich construction. The signal probes are immobilized on the surface of nanomaterials to form the signal unit, which can catalyze the reduction of a colorimetric substrate 4-nitrophenol (4-NP). Firstly, 0D gold nanoparticles (AuNPs) are employed to catalyze 4-NP reduction and reaches 102-fold signal amplification. Then AuNPs are decorated on the surface of 2D material, such as graphene oxide (GO), the catalytic efficiency is further enhanced to 104-fold signal amplification. The third step amplification is achieved by replacing stable GO with oxidizable 2D material (Bi2Se3 nanosheets), resulting in a nearly 1010-fold amplification. The sandwich-type Bi2Se3-AuNPs biosensor shows excellent sensitivity and selectivity. The detection limit can reach up to 10−18 M and there is a good linear relationship between the reaction kinetics constant and the DNA concentration in the range of 10−12-10−18 M. In addition, one-base mismatch, two-base mismatch and non-complementary sequences can be distinguished clearly by this biosensor. This design may have beneficial clinical application prospects for cancer genetic screening and early diagnosis.