CRISPR/Cas9 method is the most used method to induce targeted genomic changes. Screening and identification of CRISPR/Cas9 induced genomic changes in cell clones are expensive, time-consuming and labour intensive approaches. A high resolution melting (HRM) method enables cost-effective and rapid detection of CRISPR/Cas9 induced changes of DNA. After gDNA isolation of Chinese hamster ovarium (CHO) cells with specific reagents, we discovered that 5-fold dilution of isolated gDNA is the most optimal, based on the reproducibility of Tm value. Minimal number of cells for successful HRM method was specified, being at least 5000 CHO cells per sample. Furthermore, genomic changes of 105 clones were screened by HRM method. Every clone was targeted at two target sites inside one of five genes by CRISPR/Cas9 system. However, besides regular HRM results of five target regions, we obtained some irregular melting curves (outliers) of specific clones. Due to a considerable distance between target sites inside specific genes, we amplified those target sites with two different primer pairs. After exclusion of clones with irregular melting curves and determined them as genetic changed, we compared results of HRM method with results of next-generation sequencing method (NGS). Our results showed 95 % accuracy and 96 % sensitivity of the HRM method. If we consider clones, whose both target sites were amplified only with one pair of primer, we can conclude that we detected genetic changes with 100 % accuracy.
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