Review on the evolution in DNA-based techniques for molecular characterization and authentication of GMOs

Genetically modified (GM) foods have not only improved the yield and quality of food but also raised public concerns about the safety of GM foods. However, with the continuous innovation of sequencing technology, the characterization of GM organisms (GMOs) has been rapidly developed and upgraded several times. The traditional strategy of Southern blotting to confirm the copy number and Sanger combined with polymerase chain reaction (PCR) to identify the flanking sequences can accomplish the identification of goals, but it is not able to obtain the complete insertion information and is time-consuming and labor-intensive. Next-Generation DNA Sequencing (NGS) has effectively addressed this issue through its high-throughput sequencing technology, allowing for the rapid and cost-effective sequencing of large amounts of DNA. Its high-throughput sequencing technology has effectively addressed the limitations of traditional identification methods, offering a promising avenue for the characterization of GMOs. However, NGS is limited by the short read length and cannot identify and characterize all insertion sites included in the sequence data, especially in samples with relatively large and complex genomes. Third-generation DNA sequencing (TGS) breaks this limitation with the advantage of long reads, but its higher error rate and fewer bioinformatics tools are still the primary problems to be solved. This article presents a comprehensive review of the DNA-based techniques for assessing the molecular characterization of GM products since their introduction. The iterative evolution of these techniques is discussed, and the advantages and limitations of each method are analyzed. Through this analysis, the article identifies significant challenges and future development directions in the field of molecular characterization of GM products. By examining the strengths and weaknesses of current approaches, this review aims to provide insights and guidance for future research in this area.