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. 2022 Aug 18;4(1):vdac131. doi: 10.1093/noajnl/vdac131

Figure 2.

Figure 2

Detection of the hybrid chromosome containing the 1q/19p arms. (A) Schematic of the PCR assay that specifically detects the hybrid chromosome containing the 1q/19p arms (the 1q/19p product). To ensure reaction specificity, primer pairs recognizing 1q and 19p (arrows) and an internal Taqman probe recognizing 1q sequences have been used. INDELs, insertions and deletions of several nucleotides at the DSB repair site. Note that the internal probe is designed to recognize sequences outside the anticipated Cas9-cutting site and INDEL region. (B) Droplet digital PCR (ddPCR) analysis of the 1q/19p product in HEK 293T cells collected on different days after CRISPR/Cas9 introduction. CCR5, internal control to normalize the copy number of input genomic DNA. The percentage shows the fraction of 1q/19p positive droplets of each sample. (C) Electrophoresis of the 1q/19p product. Arrow, the expected PCR product. DNA ladder, 1KB Plus (NEB #N3200S). (D-E) Automatic sequencing analysis of the 1q/19p product using (D) the 1q primer and (E) the 19p primer. Red arrowheads, Cas9 cutting sites.