Cas9 Stable Cell Line - HEK293

The goal of the study was to improve the specificity of the CRISPR/Cas9 system for human treatments, namely in stem cell modification. They evaluated the effectiveness of current Cas9 high fidelity variants in therapeutically relevant contexts and found decreased on-target activity. They discovered a variation, HiFi Cas9 (R691A), through a bacterial screen that maintained high on-target activity with reduced off-target effects. Effective gene targeting in human CD34+ hematopoietic stem cells and primary T-cells was made possible by HiFi Cas9, including the repair of the mutation that causes sickle cell disease. This development could lead to precise genome editing for both medicinal and scientific purposes.

Figure 1. The researchers used GUIDE-seq to empirically identify off-target locations after electroporating gRNA into HEK293 cells that expressed modest amounts of either HiFi Cas9 or WT. R691A HiFi Cas9 showed a notable decrease in off-target regions while preserving a high level of on-target editing. HiFi Cas9 demonstrated an advantage in minimizing off-target events while maintaining on-target efficiency through quantitative evaluation using multiplex PCR. HiFi Cas9 consistently outperformed other high-fidelity Cas9 mutants without sacrificing on-target editing efficiency, achieving 99% or more on-target events in comparison to WT Cas9. (Vakulskas CA, et al., 2018)

Using Creative Biogene's Cas9 Stable Cell Line-HEK293 cell line can improve the above experiments to a certain extent and increase efficiency. These cell lines are optimized to provide stable Cas9 expression, reducing variability and complexity in experiments and improving the accuracy and consistency of gene editing.