The RNA-guided CRISPR-associated (Cas) nucleases are versatile tools for genome editing in various organisms. The large sizes of the commonly used Cas9 and Cas12a nucleases restrict their flexibility in therapeutic applications that use the cargo-size-limited adeno-associated virus delivery vehicle. More compact systems would thus offer more therapeutic options and functionality for this field. Here, we report a miniature class 2 type V-F CRISPR-Cas genome-editing system from Acidibacillus sulfuroxidans (AsCas12f1, 422 amino acids). AsCas12f1 is an RNA-guided endonuclease that recognizes 5′ T-rich protospacer adjacent motifs and creates staggered double-stranded breaks to target DNA. We show that AsCas12f1 functions as an effective genome-editing tool in both bacteria and human cells using various delivery methods, including plasmid, ribonucleoprotein and adeno-associated virus. The small size of AsCas12f1 offers advantages for cellular delivery, and characterizations of AsCas12f1 may facilitate engineering more compact genome-manipulation technologies.

RNA 引导的 CRISPR 相关 (Cas) 核酸酶是用于在各种生物体中进行基因组编辑的通用工具。常用的 Cas9 和 Cas12a 核酸酶的大尺寸限制了它们在使用载货尺寸受限的腺相关病毒运载工具的治疗应用中的灵活性。因此，更紧凑的系统将为该领域提供更多的治疗选择和功能。在这里，我们报告了来自Acidibacillus sulfuroxidans的微型 2 类 VF CRISPR-Cas 基因组编辑系统（AsCas12f1，422 个氨基酸）。AsCas12f1 是一种 RNA 引导的核酸内切酶，可识别富含 5' T 的原型间隔子相邻基序，并为目标 DNA 创建交错的双链断裂。我们证明 AsCas12f1 在细菌和人类细胞中作为一种有效的基因组编辑工具发挥作用，使用各种递送方法，包括质粒、核糖核蛋白和腺相关病毒。AsCas12f1 的小尺寸为细胞递送提供了优势，并且 AsCas12f1 的表征可能有助于设计更紧凑的基因组操作技术。