Transgene Mapping in Animals: What to Choose?

The phenomenal progress in biotechnology and genomics is both inspiring and overwhelming-a classic curse of choice, particularly when it comes to selecting methods for mapping transgene DNA integration sites. Transgene localization remains a crucial task for the validation of transgenic mouse or other animal models generated by pronuclear microinjection. Due to the inherently random nature of DNA integration, reliable characterization of the insertion site is essential. Over the years, a vast number of mapping methods have been developed, and new approaches continue to emerge, making the choice of the most suitable technique increasingly complex. Factors such as cost, required reagents, and the nature of the generated data require careful consideration. In this review, we provide a structured overview of current transgene mapping techniques, which we have broadly classified into three categories: classic PCR-based methods (such as inverse PCR and TAIL-PCR), next-generation sequencing with target enrichment, and long-read sequencing platforms (PacBio and Oxford Nanopore). To aid in decision-making, we include a comparative table summarizing approximate costs for the methods. While each approach has its own advantages and limitations, we highlight our top four recommended methods, which we believe offer the best balance of cost-effectiveness, reliability, and simplicity for identifying transgene integration sites.