Understanding human gene function in living organisms has long been hampered by fundamental differences between species. Although mice share most protein-coding genes with humans, their regulatory landscapes often diverge, limiting how accurately mouse models can mimic human biology. One promising solution is full-length gene humanization (FL-GH), in which entire mouse loci—including coding sequences, introns, untranslated regions, and regulatory elements—are replaced with their human counterparts. Yet existing technologies have struggled to insert very large genomic fragments efficiently or reliably, slowing efforts to develop physiologically relevant humanized models.
To address these long-standing challenges, a research team led by Associate Professor Manabu Ozawa and Associate Professor Jumpei Taguchi from The Institute of Medical Science, The University of Tokyo, Japan, has developed a streamlined two-step strategy for FL-GH. Their study, published in Nature Communications on January 14, 2026, introduces TECHNO (Two-step ES Cell-based HumaNizatiOn), a method that integrates CRISPR/Cas9-assisted genome editing with bacterial artificial chromosome (BAC)-based delivery of large human genomic regions. This framework offers a practical and scalable solution for replacing entire mouse loci with their human counterparts. "Our results demonstrate a robust and broadly applicable platform for generating FL-GH mouse models," says Dr. Ozawa.
The TECHNO workflow unfolds in two coordinated steps. First, the target mouse locus is excised using Cas9 ribonucleoproteins and replaced with short human homology arms surrounding a selection cassette, creating a precise genomic landing site. In the second step, a BAC carrying the full-length human gene and its regulatory elements is introduced into embryonic stem cells alongside a universal gRNA targeting the selection cassette, enabling homology-directed integration of genomic fragments exceeding 200 kbp. Because the method relies on standard molecular reagents and widely available BAC libraries, it is theoretically applicable to more than 90% of human genes.
Using this platform, the team successfully humanized several loci, including c-Kit, APOBEC3, and CYBB. Humanization of c-Kit reproduced human-like alternative splicing and organ-specific expression while supporting essential biological functions such as hematopoiesis and spermatogenesis. Replacement of the APOBEC3 locus demonstrated the scalability of the approach, integrating over 200 kbp of human DNA spanning seven genes and generating expression patterns that mirrored those observed in humans. The researchers also established a humanized CYBB allele and introduced disease-associated mutations to model chronic granulomatous disease. The resulting mice displayed impaired reactive oxygen species production, faithfully recapitulating the molecular phenotype found in patients.
In the near term, TECHNO is expected to accelerate the development of precise, human-relevant animal models for evaluating therapeutic targets, validating disease-associated variants, and identifying ineffective drug candidates earlier in research pipelines. Over the longer term, scalable FL-GH may reshape biomedical research by enabling models that more faithfully mimic human gene regulation and disease mechanisms. These advances also set the stage for integrating humanized models into AI-driven comparative genomics, large-scale humanized allele panels, and systems biology frameworks. As Dr. Ozawa states, "Overall, these results demonstrate that our method enables not only FL-GH of individual loci but also precise modeling of human genetic diseases in vivo by introducing disease-associated mutations into humanized alleles."
By enabling stable, high-efficiency integration of genomic fragments exceeding 200 kbp while preserving complex regulatory behavior in vivo, the TECHNO platform represents a major advance toward next-generation humanized mouse models. Its versatility, robustness, and reliance on standard laboratory tools position it as a foundational technology for advancing functional genomics, disease modeling, and translational medicine.
