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Creative Biogene CRISPR/Cas9 Platform provides CIRCLE-Seq service for customers to assess off-target effect of CRISPR/Cas9. With years of experience in CRISPR-Cas9 and NGS field, our talented scientists will work closely with you to assist you in off-target effects analysis via CIRCLE-seq.
Engineered nucleases, which can introduce targeted DNA double-stranded breaks into the genome of living cells, are widely used as genome-editing tools. However, they may introduce undesired off-target mutations as well, potentially leading to unintended effects in research and clinical applications. Since indel mutations can not be easily detected by anchored PCR amplification of an expected sequence, sensitive and unbiased genome-wide methods are developed to understand the genome locations and frequency of nuclease-induced DSB activity.
As one of the most popular methods, circularization for in vitro reporting of cleavage effects by sequencing (CIRCLE-Seq) is a novel, highly sensitive, sequencing-efficient strategy to identify CRISPR/Cas9 genome-wide off-target mutations. This assay can be practiced using widely accessible next-generation sequencing technology since it does not require reference genome sequences. Thus, CIRCLE-Seq is applied for off-target profiling in organisms lacking full genomic sequence. Comparing to Digenome-seq, CIRCLE-seq shows lower background by enrichment of nuclease-cleaved genomic DNA before sequencing. Additionally, CIRCLE-seq, as a sensitive method, can detect off-target mutations that occur with frequencies below ~0.1% in nuclease-treated cell populations.
Figure 1. Schematic overview of CIRCLE-Seq method1
CIRCLE-seq is a NGS-based method to detect off-target cleavage of CRISPR/Cas9. Genome DNA extracted from samples is sheared and circularized followed by degradation of residual linear DNA. Circular DNA molecules will be cleaved by Cas9 nuclease and then the cleavage sites are detected by high-throughput sequencing.
As a professional provider, Creative Biogene CRISPR/Cas9 Platform employs various approaches to detect the off-target cleavages of CRISPR/Cas9, especially CIRCLE-Seq. This service covers from sample preparation, sequencing to data analysis. Once project finishes, reliable report and raw data will be delivered to customers.