As a revolutionary breakthrough in the field of life sciences, genome editing technology has been able to precisely modify short DNA fragments like pruning tree branches, but editing large DNA fragments remains a challenge. On the evening of August 4, Gao Caixia's team from the Institute of Genetics and Developmental Biology of the Chinese Academy of Sciences published a paper online in the international academic journal Cell, which systematically reported a new type of programmable large segment DNA precise manipulation technology PCE at the chromosome level, realizing "precise editing" of eukaryotic genome DNA at the level of thousand base to trillion base. The reviewer commented that this work represents a significant breakthrough in the field of genetic engineering and has enormous potential for application in breeding and gene therapy. Cracking the "scale dilemma" of genome editing: DNA serves as the code of life, storing genetic information that determines biological traits, life activities, and even evolutionary directions. Currently, CRISPR and its derivative technologies, known as "gene scissors," have been widely used in specific bases and short fragments of DNA. However, precise manipulation of thousands or even millions of bases is the core challenge in editing large fragments of DNA, and existing tools still have significant shortcomings in editing efficiency, scale, accuracy, and type diversity. The research team has developed a new method for precise and traceless editing of ultra large DNA fragments, constructing two programmable chromosome editing systems, PCE and RePCE, to achieve precise and traceless manipulation of ultra large DNA fragments, successfully solving the "scale dilemma" of genome editing. Gao Caixia, a researcher at the Institute of Genetics and Developmental Biology of the Chinese Academy of Sciences, said that this technology can achieve multi type precise manipulation of millions of base level DNA in plant and animal cells, significantly improving the manipulation scale and ability of eukaryotic genomes. The Cre Lox system, a site-specific recombinase with three innovative breakthroughs, has the potential for chromosome level DNA manipulation, but its further application is constrained by three key issues. To this end, the research team has constructed a systematic technological path, achieved three key technological innovations, and installed a "navigation system" for precise and traceless editing of ultra large DNA fragments. ——Change 'two-way door' to 'single channel'. To solve the reversible problem of recombination reactions caused by the inherent symmetry of Lox sites, the research team innovatively developed a high-throughput recombination site rapid modification platform, successfully creating a new type of Lox variant. Like a one-way gate, it only allows DNA fragments to move in a predetermined direction, which is more conducive to the occurrence of targeted editing. ——Utilizing artificial intelligence to perform 'team optimization' on Cre recombinase. Simply put, Cre recombinase is a worker responsible for transporting DNA fragments. Based on the AI based protein directed evolution method AiCE independently developed by the research team, researchers have precisely optimized the Cre protein oligomerization interface, resulting in an engineered Cre protein variant with a recombination efficiency increased by 3.5 times, effectively enhancing its activity and improving "work efficiency". ——Develop a 'traceless editing strategy' for Re pegRNA. To avoid interference with the accuracy of genome editing caused by residual specific sites after recombination, the Re pegRNA strategy developed by the research team is like an intelligent eraser that can accurately identify and eliminate these residual sites, improving editing accuracy. Multi scenario applications are expected to achieve what industry insiders believe is a new method of precise and traceless editing using ultra large DNA fragments. By manipulating genomic structural variations, it can open up new paths for crop trait improvement and genetic disease treatment. In traditional breeding, excellent traits are often genetically linked to poor genes, similar to the bundled sales of "buy one get one free". The technological breakthrough of precise and traceless editing of large fragments of DNA is expected to promote the development of new breeding strategies, such as controlling fertility and eliminating linkage burdens by manipulating genetic linkage and regulating recombination frequency, fully unleashing the breeding potential of excellent alleles in wild germplasm resources. At present, the research team has successfully created a herbicide resistant rice germplasm containing precise inversion of 315 kilobases using this technology. In the field of genetic disease treatment, this technology is expected to provide new treatment ideas for diseases caused by chromosomal abnormalities. In addition, breakthroughs in precision chromosome editing technology will accelerate the construction of artificial chromosomes and have important application prospects in emerging fields such as synthetic biology. (New Society)