Black phosphorus-enhanced CRISPR/Cas9 delivery and release system for gene editing.

Recently, Yu Xuefeng, a researcher at the Shenzhen Institute of Advanced Technology of the Chinese Academy of Sciences, made a new breakthrough in the field of CRISPR gene editing.
work Enhanced Cytosolic and Delivery Releases of CRISPR/Cas9 by Phosphorus Black Nanosheets for Genome Editing (Black Phosphorus Enhanced CRISPR/Cas9 Delivery and Release System for Gene Editing) is published in German Applied Chemistry (An Gewandte Chemie International Edition, DOI: 10.1002/anie.201806941, 10.1002/ange.201806941).
the paper co-author is Dr. Zhou Wenhua and research assistant Cui Haodong, and the communications author is Yu Xuefeng, who is also involved in the author's co-author, Ying Liming, a professor at Imperial College.
In recent years, the CRISPR/Cas9 system, as a gene-targeted editing technology derived from the acquired immune system of pronuclear organisms, has been widely used in such fields as gene expression regulation, gene editing and in-situ detection of nucleic acids.
however, due to the traditional transport methods for encoded DNA, there are problems that can easily lead to the insertion loss and overexpression of target cells, and cause genetic variation and off-target effects, which limit the clinical application potential of this technology.
therefore, the development of a new high-efficiency cell transport/release method for Cas9-sgRNA nucleic acid-protein complex will have important scientific significance and application prospects.
black phosphorus has received wide attention in the biomedical field because of its good biological activity and biocompatibility, as well as its superior photoelectric properties. in previous studies, the
Yu Xuefeng team has successfully applied black phosphorus nanomaterials to photoacoustic imaging (Small, 2017, 13, 1602896), tumor photothermal therapy (Nature Communications, 2016, 7,12967), and tumor radiation therapy (Biomaterial) s, 2018, 171,12), photomorphic implants (Biomaterials, 2018, 164, 11) and hydrogel dressings (Advanced Science, 2018, 5, 1700848).
based on these early work, and the rupture of the endosome sequins caused by the degradation of black phosphorus found in these work, the team suggested that black phosphorus nanochips could be used for cell transport and release of biologically active macromolecules.
Figure 2: The (a) cytoplasmic transport of the Cas9-sgRNA complex based on black phosphorus nanochips and the degradation process within cells of nuclear transport and (b) black phosphorus nanochips.
Figure 3: Cas9-sgRNA complex transport system based on black phosphorus nanochips: (a) efficient gene editing in cell strain MCF-7; In the
study, the team used the ultra-thin two-dimensional black phosphorus nanochip as the carrier, and realized an efficient load on the Cas9-sgRNA complex carrying the three-bit sequence (3xNLS) through electrostatic interaction, thus constructing an efficient CRISPR/Cas9 gene delivery system (Figure 1).
the transport system mainly through direct membrane rupture and cytoswallowing into the cell endothelial, the black phosphorus carrier into the inner body in the acidic environment rapidly degraded to a highly biocompatible inorganic phosphate, resulting in the rupture of the internal body caused by the change of osmosis pressure, the realization of the Cas9-sgRNA complex of high-efficiency intracellular release, and finally by the three approved sequences carried into the cell nucleus, complete gene editing function (Figure 2).
experimental results show that the Cas9-sgRNA complex transfer system based on black phosphorus nanochips can perform efficient gene editing and gene silencing on different cell strains and animal tumor models at lower concentrations compared to other nanomaterial vectors (Figure 3).
this high-efficiency cell transport system based on black phosphorus nanochips can be further used for high-efficiency cell transport of other bioactive macromolecules, thus having important value in medical research and clinical application.
has applied for a patent (CN108148874A) and actively promoted the industrialization of related technologies, relying on the incubation of China Science and Phosphorus Technology Co., Ltd. (MoPhos.cn).
the research work has been supported by the Chinese Academy of Sciences' Frontier Scientific Research Focus Program, the National Natural Science Foundation, the Shenzhen Science and Technology Program International Cooperation Research, and the Lihua Hume Trust In the United Kingdom.
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