The study, published in Nature Biotechnology, describes methods using ssDNA donor templates to reduce cellular toxicity and achieve 40 percent knock-in editing efficiency with minimal cellular toxicity across a variety of primary cell types

PISCATAWAY, NJ, September 22, 2022 /PRNewswire/ — GenScript United States of America inc., the world’s leading provider of life science research tools and services, is collaborating with researchers at Gladstone-UCSF Institute of Genomic Immunology to advance the development of new, non-viral cell therapies that modify genomic sequences to create more potent cell therapy products while limiting the cellular toxicity typically associated with previously available methods.

A new study, Published in natural biotechnology, describes methods for achieving high-efficiency non-viral knock-in using GenScript’s GenExact™ single-stranded DNA (ssDNA) modified with Cas9 target sequences (CTS). This method achieved up to ~90% knock-in efficiency in primary immune cell types.

A powerful application of CRISPR/Cas genome editing technology involves the precise insertion (or knock-in) of DNA sequences via the HDR pathway. Traditionally, researchers have relied on viral vectors to deliver DNA insertion templates used for gene therapy into cells. However, difficulties in producing large quantities of clinical-grade viral vectors have delayed the delivery of cell therapies to patients. In addition, viral vectors can insert genes at any location within the genome, raising safety concerns.

Previous research by the group gladstone and UCSF have shown that synthetic DNA templates can be delivered without the use of viral vectors, but large amounts of double-stranded DNA can be toxic to cells, resulting in low efficiency. Efficiency can be improved by using a modified version of the DNA templates that can bind to the Cas9 enzyme. However, additional work was required to improve the yield of successfully constructed cells and to complete the process with the…