Current methods to delete genomic sequences are based on clustered regularly interspaced short palindromic repeats (CRISPR)–Cas9 and pairs of single-guide RNAs (sgRNAs), but can be inefficient and imprecise, with errors including small indels as well as unintended large deletions and more complex rearrangements. In the present study, we describe a prime editing-based method, PRIME-Del, which induces a deletion using a pair of prime editing sgRNAs (pegRNAs) that target opposite DNA strands, programming not only the sites that are nicked but also the outcome of the repair. PRIME-Del achieves markedly higher precision than CRISPR–Cas9 and sgRNA pairs in programming deletions up to 10 kb, with 1–30% editing efficiency. PRIME-Del can also be used to couple genomic deletions with short insertions, enabling deletions with junctions that do not fall at protospacer-adjacent motif sites. Finally, extended expression of prime editing components can substantially enhance efficiency without compromising precision. We anticipate that PRIME-Del will be broadly useful for precise, flexible programming of genomic deletions, epitope tagging and, potentially, programming genomic rearrangements.

目前删除基因组序列的方法基于成簇的规则间隔短回文重复序列 (CRISPR)–Cas9 和单向导 RNA (sgRNA) 对，但可能效率低下且不精确，错误包括小插入缺失以及意外的大删除和更复杂的重新排列。在本研究中，我们描述了一种基于初等编辑的方法 PRIME-Del，该方法使用一对针对相反 DNA 链的初等编辑 sgRNA (pegRNA) 诱导删除，不仅对切口位点进行编程，还对结果进行编程的修复。 PRIME-Del 在编程删除高达 10 kb 的过程中比 CRISPR-Cas9 和 sgRNA 对实现了明显更高的精度，编辑效率为 1-30%。 PRIME-Del 还可用于将基因组缺失与短插入结合起来，从而实现不落在原型间隔子相邻基序位点的连接处的缺失。最后，主要编辑组件的扩展表达可以在不影响精度的情况下显着提高效率。我们预计 PRIME-Del 将广泛用于精确、灵活的基因组删除编程、表位标记以及潜在的基因组重排编程。