Trans-chromosomic (Tc) mice carrying mini-chromosomes with human immunoglobulin (Ig) loci have contributed to the development of fully human therapeutic monoclonal antibodies (Abs); however, we previously observed that mitotic instability of human mini-chromosomes in mice has limited the efficiency of hybridoma production. Here, we established a new generation of human Ab producing Tc mice (TC-mAb mice), which stably maintain a mouse-derived engineered chromosome containing the entire human Ig heavy and kappa chain loci in a mouse Ig knockout background. Comprehensive, high-throughput DNA sequencing revealed that the human Ig repertoire, including variable gene use, was well recapitulated in TC-mAb mice. Despite slightly altered B cell development and a delayed immune response, immunized TC-mAb mice exhibited more subsets of antigen-specific plasmablast and plasma cells compared with wild-type mice, leading to high efficiency hybridoma production. Thus, TC-mAb mice offer a valuable platform to obtain fully human therapeutic Abs and to elucidate the regulation of human Ig repertoire formation.