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. 2024 Apr 12;22:175. doi: 10.1186/s12951-024-02427-2

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© The Author(s) 2024

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

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Fig. 4 — Validation and optimization CrLNP_6.0 on gene editing in vitro. Gene editing efficiencies by treating cells with CrLNPs targeting the surrogate reporter gene under various A lipid compositions, and B N/P ratios. C Scheme of endogenous gene (IL-10) editing in various cell lines analyzed by targeted deep sequencing. Indel frequencies of CT26 cells by treating with CrLNPs prepared at different Cas9 to sgRNA molar ratios (1:1, or 1:3) analyzed by D T7E1 assay and E NGS. F Representative sequence data from E for CrLNP_6.0 (orange: crRNA, blue: PAM, -: deletion, WT: wild-type). G Indel frequencies of various cell lines treated with CrLNP_6.0 at different molar ratios of MC3. H Changes in efficiencies of gene editing in cells treated with CrLNP_6.0 stored at 4℃ for various incubation times. I Indels at possible off-target sites in CT26 cells treated with CrLNP_6.0