Bankers Christine M, Hooper Joan E
Methods Mol Biol. 2007;397:145-60. doi: 10.1007/978-1-59745-516-9_12.
To fully understand how animals develop, it is often necessary to remove the function of a particular gene in a specific cell type or subset of cells. In Drosophila melanogaster, mosaic animals have been widely utilized to study cell fate, growth and patterning, and restriction of cell fate. This chapter describes using FLP recombinase to generate mosaic Drosophila, discussing the chromosomes and cross scheme, how to induce the clones, how to properly identify the appropriate progeny, and how to prepare and analyze the tissues, clones, and phenotypes. It then presents three examples, applying this technique to study Hedgehog signaling. The first example describes moderate-sized costal clones in imaginal discs, using green fluorescent protein (GFP) as a marker and dppLacZ and Engrailed expression as phenotypic reporters. The second describes filling the adult eye with roadkill mutant clones, using white as a marker and scoring morphology. The third describes clonal misexpression of a truncated form of Smoothened, using GFP and yellow as markers.
为了全面了解动物的发育过程，通常需要在特定细胞类型或细胞亚群中去除特定基因的功能。在黑腹果蝇中，镶嵌体动物已被广泛用于研究细胞命运、生长和模式形成以及细胞命运的限制。本章描述了使用FLP重组酶生成镶嵌体果蝇的方法，讨论了染色体和杂交方案、如何诱导克隆、如何正确鉴定合适的后代，以及如何制备和分析组织、克隆和表型。然后给出了三个例子，应用该技术研究刺猬信号通路。第一个例子描述了在成虫盘上中等大小的肋状克隆，使用绿色荧光蛋白（GFP）作为标记，dppLacZ和Engrailed表达作为表型报告基因。第二个例子描述了用roadkill突变体克隆填充成年眼睛，使用白色作为标记并对形态进行评分。第三个例子描述了使用GFP和黄色作为标记的截短形式的平滑蛋白的克隆性错误表达。
