核糖核蛋白(RNP)介导的基因组编辑是一种精确高效的基因编辑工具。利用由Cas9蛋白和导向RNA(gRNA)组成的RNP，可以实现有针对性的基因编辑，无需质粒或病毒载体。在这篇文章《Efficient peptide-mediated in vitro delivery of Cas9 RNP》中，Gustafsson等人(2021)描述了一种高效的肽介导的体外Cas9 RNP传递方法。

将Cas9 RNP传递至细胞内是基因组编辑的关键步骤。传统的方法，如电穿孔或转染，效率低且可能导致细胞毒性。肽介导传递是一种具有吸引力的替代方法，因为它无毒且高效。在他们的研究中，Gustafsson等人使用一种叫做PepFect14的肽将Cas9 RNP传递至细胞内。

PepFect14是一种能够高效传递核酸至细胞内的穿膜肽。在他们的研究中，Gustafsson等人将PepFect14与Cas9蛋白和gRNA偶联，形成Cas9 RNP复合物。然后，他们使用HEK293T细胞体外测试了这种传递方法的效率。他们发现，PepFect14介导的Cas9 RNP传递可以高效进行基因编辑，同时细胞毒性较小。

与传统方法相比，使用PepFect14进行Cas9 RNP传递具有几个优势。首先，它无毒，不需要使用病毒载体或质粒。这减少了非特异性效应的风险，并消除了耗时的克隆步骤的需求。其次，PepFect14介导的传递效率非常高，可以实现强大的基因编辑且细胞死亡较少。最后，PepFect14是一种商业可获得的试剂，使其全球研究人员都可以使用。

除了PepFect14，还有其他肽已用于Cas9 RNP的传递。例如，一种叫做NickFect的肽已被

证明能够高效将Cas9 RNP传递至细胞内。NickFect是一种阳离子肽，能够与Cas9 RNP形成复合物，实现高效传递至细胞内。另一种叫做Cas9-PEG的肽则能够提高Cas9 RNP的稳定性和传递性。Cas9-PEG是一种与聚乙二醇结合的肽，能够保护Cas9 RNP免受降解并改善其传递至细胞内。

与传统方法相比，使用RNP进行基因组编辑具有几个优势。首先，RNP可以直接传递至细胞内，实现精确高效的基因编辑，无需质粒或病毒载体。其次，RNP的半衰期较短，比质粒或病毒载体低，从而降低了非特异性效应的风险。最后，RNP可以轻松修饰以针对特定基因，实现精确的基因编辑。

总之，使用RNP进行基因组编辑是一种精确高效的基因编辑工具。使用肽，如PepFect14，进行RNP传递是传统方法的一种有吸引力的替代方案，因为它无毒且高效。新肽，如NickFect和Cas9-PEG的开发，进一步扩大了RNP传递的选择。使用RNP进行基因组编辑有潜力革命化基因治疗和个性化医学领域。

References:

1.

2. Lee, K., Conboy, M., Park, H. M., Jiang, F., Kim, H. J., Dewitt, M. A., ... & Murthy, N. (2017).  Nature Biomedical Engineering, 1(11), 889-901.

Ribonucleoprotein (RNP)-mediated genome editing is a powerful tool for precise and efficient gene editing. The use of RNPs, which consist of the Cas9 protein and a guide RNA (gRNA), allows for targeted gene editing without the need for plasmids or viral vectors. In this article, 《Efficient peptide-mediated in vitro delivery of Cas9 RNP. Pharmaceutics》.Gustafsson et al. (2021) describe an efficient peptide-mediated in vitro delivery method for Cas9 RNPs.

The delivery of Cas9 RNPs into cells is a critical step in genome editing. Traditional methods, such as electroporation or transfection, can be inefficient and may cause cellular toxicity. Peptide-mediated delivery is an attractive alternative, as it is non-toxic and can be highly efficient. In their study, Gustafsson et al. used a peptide called PepFect14 to deliver Cas9 RNPs into cells.

PepFect14 is a cell-penetrating peptide that can efficiently deliver nucleic acids into cells. In their study, Gustafsson et al. conjugated PepFect14 to the Cas9 protein and gRNA to form a Cas9 RNP complex. They then tested the efficiency of this delivery method in vitro using HEK293T cells. They found that PepFect14-mediated delivery of Cas9 RNPs resulted in efficient gene editing with minimal cellular toxicity.

The use of PepFect14 for Cas9 RNP delivery has several advantages over traditional methods. First, it is non-toxic and does not require the use of viral vectors or plasmids. This reduces the risk of off-target effects and eliminates the need for time-consuming cloning steps. Second, PepFect14-mediated delivery is highly efficient, allowing for robust gene editing with minimal cell death. Finally, PepFect14 is a commercially available reagent, making it accessible to researchers worldwide.

In addition to PepFect14, other peptides have been used for Cas9 RNP delivery. For example, a peptide called NickFect has been shown to efficiently deliver Cas9 RNPs into cells. NickFect is a cationic peptide that forms a complex with Cas9 RNPs, allowing for efficient delivery into cells. Another peptide, called Cas9-PEG, has been shown to improve the stability and delivery of Cas9 RNPs. Cas9-PEG is a polyethylene glycol-conjugated peptide that can protect Cas9 RNPs from degradation and improve their delivery into cells.

The use of RNPs for genome editing has several advantages over traditional methods. First, RNPs can be delivered directly into cells, allowing for precise and efficient gene editing without the need for plasmids or viral vectors. Second, RNPs have a shorter half-life than plasmids or viral vectors, reducing the risk of off-target effects. Finally, RNPs can be easily modified to target specific genes, allowing for precise gene editing.

In conclusion, the use of RNPs for genome editing is a powerful tool for precise and efficient gene editing. The use of peptides, such as PepFect14, for RNP delivery is an attractive alternative to traditional methods, as it is non-toxic and highly efficient. The development of new peptides, such as NickFect and Cas9-PEG, further expands the options for RNP delivery. The use of RNPs for genome editing has the potential to revolutionize the field of gene therapy and personalized medicine.

References:

1. Wang, M., Glass, Z. A., Xu, Q., & Zhu, J. (2019). Efficient delivery of genome-editing proteins using bioreducible lipid nanoparticles. Journal of Biological Chemistry, 294(29), 11343-11350.

2. Lee, K., Conboy, M., Park, H. M., Jiang, F., Kim, H. J., Dewitt, M. A., ... & Murthy, N. (2017). Nanoparticle delivery of Cas9 ribonucleoprotein and donor DNA in vivo induces homology-directed DNA repair. Nature Biomedical Engineering, 1(11), 889-901.

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