Deletion and replacement of long genomic sequences using prime editing
RNA Therapeutics Institute; Program in Bioinformatics and Integrative Biology; Li Weibo Institute for Rare Diseases Research
Bioinformatics | Genetics and Genomics
Genomic insertions, duplications and insertion/deletions (indels), which account for ~14% of human pathogenic mutations, cannot be accurately or efficiently corrected by current gene-editing methods, especially those that involve larger alterations (>100 base pairs (bp)). Here, we optimize prime editing (PE) tools for creating precise genomic deletions and direct the replacement of a genomic fragment ranging from ~1 kilobases (kb) to ~10 kb with a desired sequence (up to 60 bp) in the absence of an exogenous DNA template. By conjugating Cas9 nuclease to reverse transcriptase (PE-Cas9) and combining it with two PE guide RNAs (pegRNAs) targeting complementary DNA strands, we achieve precise and specific deletion and repair of target sequences via using this PE-Cas9-based deletion and repair (PEDAR) method. PEDAR outperformed other genome-editing methods in a reporter system and at endogenous loci, efficiently creating large and precise genomic alterations. In a mouse model of tyrosinemia, PEDAR removed a 1.38-kb pathogenic insertion within the Fah gene and precisely repaired the deletion junction to restore FAH expression in liver.
Genetic engineering, Targeted gene repair
DOI of Published Version
Jiang T, Zhang XO, Weng Z, Xue W. Deletion and replacement of long genomic sequences using prime editing. Nat Biotechnol. 2021 Oct 14. doi: 10.1038/s41587-021-01026-y. Epub ahead of print. PMID: 34650270. Link to article on publisher's site
Jiang T, Zhang X, Weng Z, Xue W. (2021). Deletion and replacement of long genomic sequences using prime editing. UMass Chan Medical School Faculty Publications. https://doi.org/10.1038/s41587-021-01026-y. Retrieved from https://escholarship.umassmed.edu/faculty_pubs/2165