The purpose of this study is to demonstrate the effects of biodiesel fraction on auto-ignition for gasoline–biodiesel blended fuel, which combines two fuels with widely different auto-ignition characteristics. First, gasoline was blended with biodiesel at 5%, 10%, 15%, and 20% by volume, and then tested in a rapid compression expansion machine at a compression ratio of 11 and a temperature range of 720–850 K to observe the auto-ignition delay phenomenon under engine-like conditions. The experimental conditions are focused on improving the auto-ignition characteristic of gasoline direct-injection compression ignition combustion strategies under low load and cold start. The heat release rate of the blended fuels was calculated from the pressure trace and displacement history of the piston in order to identify first-stage ignition and second-stage (auto-ignition) ignition delay. Second, a gasoline–biodiesel reaction mechanism was developed to predict the chemical ignition delay of the blended fuels. The reaction mechanism with 4285 species and 15,246 reactions was validated and implemented using the CHEMKIN PRO software. Finally, the chemical ignition delay was predicted by the simulation which was further compared to the experimental measured results. These results revealed that a higher biodiesel fraction helps to obtain shorter ignition delay, which reduces the requirement of intake temperature. The blended fuel with 20% biodiesel showed the lowest ambient temperature at the injection timing requirement and was 80 K lower than gasoline. Each blended fuel exhibited two-stage ignitions in the measured temperature range. The combustion duration and pressure peak of every blended fuel were similar to each other after increasing the biodiesel fraction.