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Due to their substantial volume, subsoils contain more of the total soil carbon (C) pool than topsoils. Much of this C is thousands of years old, suggesting that subsoils offer considerable potential for long-term C sequestration. However, knowledge of subsoil C behaviour and manageability remains incomplete, and subsoil C storage potential has yet to be realised at a large scale, particularly in agricultural systems. A range of biological (e.g. deep-rooting), chemical (e.g. biochar burial) and physical (e.g. deep ploughing) C sequestration strategies have been proposed, but are yet to be assessed. In this review, we identify the main factors that regulate subsoil C cycling and critically evaluate the evidence and mechanistic basis of subsoil strategies designed to promote greater C storage, with particular emphasis on agroecosystems. We assess the barriers and opportunities for the implementation of strategies to enhance subsoil C sequestration and identify 5 key current gaps in scientific understanding. We conclude that subsoils, while highly heterogeneous, are in many cases more suited to long-term C sequestration than topsoils. The proposed strategies may also bring other tangible benefits to cropping systems (e.g. enhanced water holding capacity and nutrient use efficiency). Furthermore, while the subsoil C sequestration strategies we reviewed have large potential, more long-term studies are needed across a diverse range of soils and climates, in conjunction with chronosequence and space-for-time substitutions. Also, it is vital that subsoils are more consistently included in modelled estimations of soil C stocks and C sequestration potential, and that subsoil-explicit C models are developed to specifically reflect subsoil processes. Finally, further mapping of subsoil C is needed in specific regions (e.g. in the Middle East, Eastern Europe, South and Central America, South Asia and Africa). Conducting both immediate and long-term subsoil C studies will fill the knowledge gaps to devise appropriate soil C sequestration strategies and policies to help in the global fight against climate change and decline in soil quality. In conclusion, our evidence-based analysis reveals that subsoils offer an untapped potential to enhance global C storage in terrestrial ecosystems. • Agricultural subsoils may be more suited to long-term C sequestration than topsoils. • Sequestration strategies have variable effects on C stocks, depending on soil type. • Enhanced subsoil C storage can bring ancillary tangible benefits to cropping systems. • Subsoil sampling and incorporation in models and maps is essential. • Policies are needed for C sequestration in the whole soil profile, not just topsoils. [ABSTRACT FROM AUTHOR]