Early embryonic development is driven exclusively by maternal gene products deposited into the oocyte. Although critical in establishing early developmental programs, maternal gene functions have remained elusive due to a paucity of techniques for their systematic disruption and assessment. CRISPR-Cas13 systems have recently been employed to degrade RNA in yeast, plants, and mammalian cell lines. However, no systematic study of the potential of Cas13 has been carried out in an animal system. Here, we show that CRISPR-RfxCas13d (CasRx) is an effective and precise system to deplete specific mRNA transcripts in zebrafish embryos. We demonstrate that zygotically expressed and maternally provided transcripts are efficiently targeted, resulting in a 76% average decrease in transcript levels and recapitulation of well-known embryonic phenotypes. Moreover, we show that this system can be used in medaka, killifish, and mouse embryos. Altogether, our results demonstrate that CRISPR-RfxCas13d is an efficient knockdown platform to interrogate gene function in animal embryos.

早期胚胎发育完全由沉积在卵母细胞中的母体基因产物驱动。尽管在建立早期发育计划中至关重要，但由于缺乏系统的干扰和评估技术，孕产妇的基因功能仍然难以捉摸。CRISPR-Cas13系统最近已被用于降解酵母，植物和哺乳动物细胞系中的RNA。但是，尚未在动物系统中对Cas13的潜力进行系统的研究。在这里，我们证明了CRISPR-RfxCas13d（CasRx）是一种有效且精确的系统，可以消除斑马鱼胚胎中特定的mRNA转录物。我们证明，合子表达和母本提供的成绩单是有效的针对性，导致成绩单水平平均下降76％和知名胚胎表型的概括。此外，我们证明了该系统可用于青aka，致死鱼和小鼠胚胎。总之，我们的结果表明，CRISPR-RfxCas13d是一个有效的敲低平台，可在动物胚胎中检验基因功能。