Environmental issues have become of crucial importance in the transport sector. Transport is the second biggest greenhouse gas emitting sector after energy and is responsible for 25% of the EU's total emissions. The challenges posed by climate change have added to the urgency for developing low-carbon transportation. In this paper, estimation of greenhouse gas emissions was conducted over the construction and the operation of the main road and rail axes infrastructure in Greece. The objective of this analysis is to better understand the significance of these emissions and their possible influence on designing optimal routes in order to achieve long-term greenhouse gas reductions from transport. The present study shows that the environmental impact due to the highway construction is smaller than that of the railway construction. However, the railway system operation is more environmentally friendly than the highway operation.