Cas9 screen技术是一项极具潜力的高通量筛选工具,已被应用于各个领域。同时,其衍生与拓展技术,如Cas13 screen、Base editor screen也逐步问世。我相信未来的研究手段一定是从杂乱走向整合、从非系统性走向系统性、从低通量走向高通量的,因此类似Cas9 screen的技术在未来一定大有可为。 参考 ^Shalem, Ophir et al. ...
目前表型分析从简单的增值到复杂的药物抗性、迁移和标记物的基因表达等等。 CRISPR-Cas9 文库高通量筛选常用模式可以分为阳性筛选 (Positive selection screen) 和阴性筛选 (Negative selection screen)。阳性筛选是对已成功整合 sgRNA 的细胞文库施加一定的筛选压力,仅使少数目的表型的细胞能够存活,达到富集关键基因的目的...
Thus, we conducted an additional CRISPR-Cas9 knockout screen under 5-AzaC treatment or corresponding DMSO control in SMB21 cells, observing good screening signal as assessed by the depletion of known pan-essential genes in the DMSO arm (Additional file 1: Fig. S7a). In our knockout screen ...
Figure 1. CRISRP/Cas9 Screens Identify Growth and Survival Factors in EBV-Infected Burkitt and Lymphoblastoid B Cells (A) CRISPR/Cas9 screen workflow and screening strategy. (B and C) Scatterplots showing the statistical significance of top (B) GM12878-selective and (C) P3HR1-selective hits....
Genome-wide functional screening using the CRISPR-Cas9 system is a powerful tool to uncover tumor-specific and common genetic dependencies across cancer cell lines. Current CRISPR-Cas9 knockout libraries, however, primarily target protein-coding genes. T
Lentiviral transduction of Cas9-expressing cells with LentiPool sgRNA library Selection with puromycin Expansion of resistant cells Primary screening by positive or negative selection: Positive selection (+): Apply treatment to cells (e.g., drugs, chemical perturbant) Negative selection (–): Divide...
We combined such negative selection pressure with ccdB plasmid-based positive selection pressure to develop “Sniper-screen,” which selects for Cas9 variants with increased specificities. Note that Cas9 variants with poor on-target activities or poor specificities cannot survive in this selection ...
Lentiviral transduction of Cas9-expressing cells with LentiPool sgRNA library Selection with puromycin Expansion of resistant cells Primary screening by positive or negative selection: Positive selection (+): Apply treatment to cells (e.g., drugs, chemical perturbant) Negative...
自2014年张锋等首次将CRISPR/Cas9应用于大规模基因筛选以来,CRISPR Screen 文库作为一个高效的工具,已成功地应用于包括多种细胞系及人、小鼠等模式动物中,参与功能基因筛选、药物和耐药靶点筛选及信号通路揭示等。 2.选择双载体文库还是单载体? 问题1:很多老师会问这两个的区别究竟是什么?
We therefore aimed to identify novel epigenetic treatment targets in CSC derived from H3-mutated high-grade glioma (HGG) through a CRISPR-Cas9 knockout screen.#The knockout screen identified more than 100 novel genes essential for the growth of CSC derived from paediatric HGG with H3K27M ...