More than 20 different PIK3CA gene mutations were identified in breast cancer with different frequencies. Whether these breast cancer associated mutations have similar biological effects is largely unknown. In this study, we established a novel cell model using the lentivirus system to express 10 different PIK3CA genes (wild type and mutant) based on the human mammary epithelial cell MCF10A. We found that nine different PIK3CA mutants harbor different abilities to promote cell proliferation and EGF independent growth. In addition, most PIK3CA mutants (except for the wild type PIK3CA, the Q60K and the K111N mutants) had the ability to change the morphogenesis of the MCF10A cell in 3D Matrigel assay. Moreover, different PIK3CA mutants have different abilities to promote colony formation and cell invasion. We further observed that most of the PIK3CA mutants could activate p-AKT and p-p70-S6K in the absence of EGF stimulation. Finally, LY294002, a PI3K inhibitor, can effectively inhibit cell growth in cell lines with different PIK3CAs. Taken together, our results support the notion that different PIK3CA mutations differentially contribute to breast cancer transformation, and exploration of the therapeutic application of these mutations will benefit breast cancer patients with the PIK3CA mutations.