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DNA-guided genome editing using the Natronobacterium gregoryi Argonaute

Nature Biotechnology volume 34, pages 768773 (2016) | Download Citation

  • An Addendum to this article was published on 28 November 2016
  • This article was retracted on 08 August 2017

This article has been updated


The RNA-guided endonuclease Cas9 has made genome editing a widely accessible technique. Similar to Cas9, endonucleases from the Argonaute protein family also use oligonucleotides as guides to degrade invasive genomes. Here we report that the Natronobacterium gregoryi Argonaute (NgAgo) is a DNA-guided endonuclease suitable for genome editing in human cells. NgAgo binds 5′ phosphorylated single-stranded guide DNA (gDNA) of 24 nucleotides, efficiently creates site-specific DNA double-strand breaks when loaded with the gDNA. The NgAgo–gDNA system does not require a protospacer-adjacent motif (PAM), as does Cas9, and preliminary characterization suggests a low tolerance to guide–target mismatches and high efficiency in editing (G+C)-rich genomic targets.

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  • 28 November 2016

    The editors of Nature Biotechnology are issuing an editorial expression of concern regarding this article to alert our readers to concerns regarding the reproducibility of the original results. At this time, we are publishing the results of three groups ( that have tried to reproduce the results in the critical Figure 4 in the original paper by Han and colleagues, which demonstrates editing of endogenous genomic loci in mammalian cells. None of the groups observed any induction of mutations by NgAgo at any of the loci or under any of the conditions tested above the sensitivity of the assays used. Similar results have been recently reported by a different group of authors in Protein & Cell (doi:10.1007/s13238-016-0343-9).  We are in contact with the authors, who are investigating potential causes for the lack of reproducibility. The authors have been informed of this statement. While the investigations are ongoing, Chunyu Han and Xiao Z. Shen agree with this editorial expression of concern. Feng Gao, Feng Jiang and Yongqiang Wu do not feel that it is appropriate at this time.  We will update our readers once these investigations are complete.

  • 02 August 2017

    We are retracting our study because of the continued inability of the research community to replicate the key results in Figure 4, using the protocols provided in our paper. In this figure we report that the Natronobacterium gregoryi Argonaute can efficiently create double-strand breaks and edit the genome of human cells using 5ʹ phosphorylated single-stranded DNA as a guide. Despite the efforts of many laboratories (Protein Cell 7, 913–915, 2016; Nat. Biotechnol. 35, 17–18, 2017; Cell Res. 26, 1349–1352, 2016; PLOS One 12, e0177444, 2017), an independent replication of these results has not been reported. We are therefore retracting our initial report at this time to maintain the integrity of the scientific record. We nevertheless continue to investigate the reasons for this lack of reproducibility with the aim of providing an optimized protocol.


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We thank W. Chao for performing flow cytometry experiment. This work was supported by the National Science Foundation of China 31270950 to X.Z.S.

Author information


  1. Department of Bioscience and Bioengineering, Hebei University of Science and Technology, Shijiazhuang, Hebei, China.

    • Feng Gao
    • , Feng Jiang
    • , Yongqiang Wu
    •  & Chunyu Han
  2. Department of Physiology, Zhejiang University School of Medicine, Hangzhou, Zhejiang, China.

    • Xiao Z Shen


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C.H. conceived the study and designed the experiments. X.Z.S. provided intellectual advice on the project and experimental design. C.H. performed mammalian genome editing. F.G. performed the BLAST search and the in vitro cleavage experiments. F.G., F.J. and Y.W. designed and constructed the clones under the supervision of C.H., F.J. and Y.W. performed the in vivo cleavage experiments. X.Z.S. and C.H. wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Chunyu Han.

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