NgAgo-gDNA technology is a gene editing technology that uses DNA as a guidance tool. The working principle of NgAgo-gDNA technology is somewhat similar to that of CRISPR-Cas9 technology. Under the guidance of the guidance tool, the nuclease cuts the gene sequence at a specific site to perform gene editing. The difference is that the guidance tool used in NgAgo-gDNA technology is a guide DNA (gDNA) instead of RNA in CRISPR-Cas9 technology. Since there is no need to use proteins (such as zinc finger proteins) to find sequences that need to be replaced, NgAgo-gDNA technology, like CRISPR-Cas9 technology, is much simpler and more convenient to operate than previous gene editing technologies, which is beneficial to its use. promotion in applications.
The nuclease used in NgAgo-gDNA technology is NgAgo, an Ago endonuclease protein found in Natronobacterium gregoryi. Ago nuclease was originally discovered by Dutch scientists that it can effectively use single-stranded DNA as a short medium to cleave genomic targets relatively accurately. The limitation of the initial research was that the temperature required for the experiment was 65-75 degrees Celsius, which could not be completed under physiological conditions. Through continuous searches by Professor Han Chunyu's team, they finally discovered that the Ago homologous protein from Halobacterium gastrophila can achieve similar functions under physiological conditions.
NgAgo-gDNA technology may have more advantages than CRISPR-Cas9 technology. Compared with CRISPR-Cas9 technology, NgAgo-gDNA technology has a wider selection of target sites that can be edited. Because Cas9 needs to pair with 19 bases on the genome and requires a specific three-base sequence (PAM sequence) immediately after this set of bases, it limits the selection of target sites to a certain extent, and NgAgo-gDNA technology The selection of the target site is not limited by the PAM sequence, and the editing objects are less restricted, and can edit almost any position in the genome.
In addition, the length of gDNA bound to NgAgo is 24 bases, which is 5 bases longer than the 19 bases gRNA bound to Cas9. Theoretically, its accuracy should be improved by 1024 (4 5th power) times. Moreover, research by Han Chunyu's team also found that compared with CRISPR-Cas9, the NgAgo-gDNA system has a very low tolerance for guide sequence-target sequence mismatches. The editing accuracy is higher and can more effectively avoid off-target phenomena.