The growth of worldwide environmental awareness has prompted numerous countries to focus on developing energy-conservation and carbon-reduction technology. The advancement of such technology enables the emergence of low-noise, low-polluting alternatives for bus systems, such as hybrid-electric, battery-electric, and fuel-cell electric buses (e-buses). For such buses to serve the existing schedules and lines operated by their conventional counterparts, reorganizing bus transportation systems is a major challenge and entails construction costs that comprise the costs of e-buses, battery capacity, chargers, and bus scheduling. To facilitate the development of environmentally friendly public transportation, this study proposes a model for simulating the operation and battery charging schedule of plug-in e-buses on the basis of an existing schedule and line network. The model was used to estimate the overall construction cost of converting the existent bus transportation system into an all plug-in e-bus one. Focusing on the bus transportation system in Penghu, an archipelago of Taiwan, this case study examined the effects of day- and nighttime charging requirements on the construction cost of an e-bus transportation system to improve the practicability of e-buses. It also applied a genetic algorithm to determine the minimum construction cost, which varied depending on the number of e-buses, level of battery capacity, number of chargers, and electricity costs. The optimized parameters involved the hourly residual battery capacity and battery charging times during the daytime operating hours. The results showed that although daytime charging involved electricity uses during peak hours and thus incurred additional costs, it contributed to the use of e-buses and an overall reduction in the construction cost. In summary, the proposed optimization method would successfully reduce the construction cost of the Penghu e-bus transportation system.