Chinese scientists develop new gene editing technology to replace Cas9
On May 2, in a study published in Nature Biotechnology, researchers at Hebei University of Science and Technology and Zhejiang University School of Medicine reported that Natronobacterium gregoryi Argonaute (NgAgo) is a DNA-directed nucleic acid that can be used for human cell gene editing. Endonuclease.
RNA-directed endonuclease Cas9 makes gene editing a widely used technique. However, although scientists have tried various ways to optimize the Cas9 system to improve its efficiency and specificity, the practicality of the system is still limited by some factors, including its guide-targeting. The tolerance of mismatches and the targeting of RNA are relatively easy to form secondary structures and the like.
Similar to Cas9, endonucleases from the Argonaute protein family can also use oligonucleotides as a guide to degrade invasive genomes. Argonautes plays a key role in gene expression inhibition and defense against foreign nucleic acids. In a study published on PNAS on April 12, researchers at the University of California at Berkeley revealed an atypical Argonaute-directed RNA specificity. The paper was co-authored by Jennifer A. Doudna, one of the pioneers of CRISPR.
On May 2, the latest study published in Nature Biotechnology, researchers at Hebei University of Science and Technology and Zhejiang University School of Medicine reported that Natronobacterium gregoryi Argonaute (NgAgo) is a DNA-directed gene editing for human cells. Endonuclease. NgAgo binds to a 5' phosphorylated single-stranded guide DNA (gDNA) of approximately 24 nucleotides in size, effectively forming a site-specific DNA double-strand gap.
It is reported that the targeting specificity of CRISPR/Cas9 is determined by two parts, one is the base pairing between RNA chimera and target DNA, and the other is the combination of Cas9 protein and a short DNA motif. This short DNA motif is usually found at the 3' end of the target DNA and is called the protospacer adjacent motif (PAM).
Unlike Cas9, the NgAgo–gDNA system does not require PAM, and preliminary identification indicates that the system is less tolerant to guide-target mismatches and more effective in editing G+C-rich genomes. According to reports, Cas9 is only found in prokaryotes, and Argonautes is found in almost all organisms.
In addition, in order to properly bind to Cas9, the targeting RNA must have a 3' RNA-RNA hybrid structure, and binding to Argonaute does not require any specific secondary structure of the targeting molecule. On the other hand, Cas9 can only cleave sequences upstream of the PAM, while Argonaute does not require specific sequences for the target. The authors say that Natronobacterium gregoryi Argonaute is expected to be a precise and effective tool for editing mammalian genomes.
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