Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York, United States of America.
Weill Institute for Cell and Molecular Biology, Cornell University, Ithaca, New York, United States of America.
PLoS Biol. 2021 Jan 14;19(1):e3001061. doi: 10.1371/journal.pbio.3001061. eCollection 2021 Jan.
Mosaic animals have provided the platform for many fundamental discoveries in developmental biology, cell biology, and other fields. Techniques to produce mosaic animals by mitotic recombination have been extensively developed in Drosophila melanogaster but are less common for other laboratory organisms. Here, we report mosaic analysis by gRNA-induced crossing-over (MAGIC), a new technique for generating mosaic animals based on DNA double-strand breaks produced by CRISPR/Cas9. MAGIC efficiently produces mosaic clones in both somatic tissues and the germline of Drosophila. Further, by developing a MAGIC toolkit for 1 chromosome arm, we demonstrate the method's application in characterizing gene function in neural development and in generating fluorescently marked clones in wild-derived Drosophila strains. Eliminating the need to introduce recombinase-recognition sites into the genome, this simple and versatile system simplifies mosaic analysis in Drosophila and can in principle be applied in any organism that is compatible with CRISPR/Cas9.
嵌合体动物为发育生物学、细胞生物学和其他领域的许多基础发现提供了平台。通过有丝分裂重组产生嵌合体动物的技术在黑腹果蝇中得到了广泛的发展，但在其他实验室生物中则不太常见。在这里，我们报告了基于 CRISPR/Cas9 产生的 DNA 双链断裂的 gRNA 诱导的交叉（MAGIC）的嵌合体分析，这是一种基于 CRISPR/Cas9 产生嵌合体动物的新技术。MAGIC 可有效地在果蝇的体组织和生殖系中产生嵌合体克隆。此外，通过开发用于 1 条染色体臂的 MAGIC 工具包，我们证明了该方法在神经发育中表征基因功能以及在野生衍生的果蝇品系中产生荧光标记克隆中的应用。该方法通过消除在基因组中引入重组酶识别位点的需要，简化了果蝇中的嵌合体分析，并且原则上可以应用于任何与 CRISPR/Cas9 兼容的生物体。
