Reference genomes of root microbes are essential for metagenomic analyses and mechanistic studies of crop root microbiomes. Combining high-throughput bacterial cultivation with metagenomic sequencing, we constructed comprehensive bacterial and viral genome collections from the roots of wheat, rice, maize, and Medicago. The crop root bacterial genome collection (CRBC) significantly expands the quantity and phylogenetic diversity of publicly available crop root bacterial genomes, with 6,699 bacterial genomes (68.9 % from isolates) and 1,817 novel species, expanding crop root bacterial diversity by 290.6%. The crop root viral genome collection (CRVC) contains 9,736 nonredundant viral genomes, with 1,572 previously unreported genus-level clusters in crop roots. From these data, we identified conserved bacterial functions enriched in root microbiomes across soils and host species and uncovered previously unexplored bacteria–virus connections in crop root ecosystems. Together, the CRBC and CRVC serve as valuable resources for investigating microbial mechanisms and applications, supporting sustainable agriculture. The gene sets of 14 datasets are available under this accession.