Limiting Off-Target Effects Of CRISPR-Based Products: Guide RNA Sequencing By NGS

By Rebecca A. Bova , Ph.D., Christina Kalinyak, Maggie Luce, Brianna McManus, Servane Orozco, and Bradley Hasson

In this poster, a groundbreaking study conducted by researchers at MilliporeSigma, advancements in CRISPR-Cas9 gene editing technologies have been made through enhanced quality control measures for single-guide RNA (sgRNA) using Next Generation Sequencing (NGS). This research is particularly timely, following the FDA's approval of the first CRISPR-Cas9 product in late 2023, underscoring the increasing significance of CRISPR-based applications. The study addresses a critical concern: the specificity of sgRNA is vital to ensure accurate targeting of genomic locations, as errors can result in unintended "off-target" edits. Traditional quality control methods, such as mass spectrometry, often fail to detect subtle sequence variations, prompting the need for a more sensitive approach. The researchers introduced a novel method utilizing short-read NGS technology, successfully evaluating various sgRNA kits from multiple manufacturers.

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