Genome edit detection (CRISPR-Cas9)

Nucleases such as zinc-finger (ZFN), transcription activator-like effector (TALEN), and clustered regularly interspaced short palindromic repeat (CRISPR) are used to edit the genome of any cell. These nucleases produce site-specific DNA double-strand breaks (DSBs), which then can be repaired by imprecise, error-prone non-homologous end joining (NHEJ) (donor template/precise point mutation) or by homology-directed repair (HDR) (deletion/indels/insertions) pathways leading to targeted mutagenesis. As a result, a mixed population of cells with heterogeneous indel errors and varying allelic editing frequencies develop. Then, genome editing frequencies at the desired locus are measured. Clonal cell lines isolate single cells, which are then assayed to verify the genome editing event.

Benefits of using nanoplate dPCR for genome edit detection (CRISPR-Cas9)

• Higher sensitivity enabling detection of editing events present at frequencies of 0.5%
• Absolute quantification of editing events from as little as 5 ng of total gDNA
• Ability to distinguish between homozygous and heterozygous edits in clonal populations

Related publications