CRISPR/Cas9-mediated genome editing technology consists of a single-guide RNA (sgRNA), and the Cas9 endonuclease has the potential to treat genetic diseases in most tissues and organisms. In this system, the Cas9 protein can be directed to target genomic DNA sequences as “molecular scissors” with the guidance of sgRNAs. However, the target-specific activities of different sgRNAs are highly variable; thus, it is crucial to search for a simple, quick and economical method to screen for optimized sgRNAs with high target specificity. We have adopted and verified a newly developed white-to-blue colony formation assay to quickly screen for sgRNAs optimized for the EphA2 gene, which is highly expressed in hormone-resistant prostate cancer (PC-3) cells. This assay promises to screen for optimized sgRNAs more simply, rapidly, and efficiently. Our results suggest that the white-to-blue colony formation assay might be a useful screening strategy to quickly select for optimized sgRNAs.

CRISPR/ Cas9介导的基因组编辑技术由单引导RNA（sgRNA）组成， Cas9核酸内切酶具有治疗大多数组织和生物体遗传疾病的潜力。在该系统中， Cas9蛋白可以在sgRNA的引导下像“分子剪刀”一样定向到目标基因组DNA序列。然而，不同 sgRNA 的靶标特异性活性差异很大；因此，寻找一种简单、快速、经济的方法来筛选具有高靶点特异性的优化sgRNA至关重要。我们采用并验证了一种新开发的白到蓝集落形成测定法，可快速筛选针对EphA2基因优化的 sgRNA，该基因在激素抵抗性前列腺癌 (PC-3) 细胞中高表达。该测定有望更简单、更快速、更高效地筛选优化的 sgRNA。我们的结果表明，白到蓝集落形成测定可能是一种有用的筛选策略，可快速选择优化的 sgRNA。