Expanding public transportation and conserving biodiversity are two major components of the United Nations Sustainable Development Goals. However, these objectives often conflict, as habitat loss and fragmentation caused by transportation networks and subsequent development are regarded as significant threats to biodiversity. Quantifying the extent of these contradictions and identifying strategies for balancing these goals are essential. In this study, an index of natural habitat areas without roads (i.e., roadless areas) is employed to quantify the contradiction and analyze changes in China's roadless areas from 2000 to 2020, with projections for potential changes from 2020 to 2035. The findings indicate that China's roadless area decreased significantly from 4.94 million km2 in 2000 to 4.31 million km2 in 2020, reflecting a reduction rate of 12.7%. It is conservatively estimated that this loss will continue at a rate of 4.58% over the next 15 years. Currently, only 22.8% and 35.8% of the total roadless area in 2020 are situated within biodiversity conservation priority areas and protected areas, respectively, highlighting significant conservation gaps. To address the rapid decline and inadequate protection of roadless areas in China, this study proposes several recommendations, including optimizing transportation route planning to avoid unnecessary loss of roadless area, constructing wildlife crossing structures to enhance habitat connectivity, and integrating high ecological value roadless areas into the conservation framework. This study offers valuable insights into the effective protection of ecosystem integrity amidst the expansion of road networks and presents new solutions for ensuring timely compliance with international agreements, such as the Convention on Biological Diversity.