Mutant analysis by rescue gene excision: New tools for mosaic studies in Drosophila.
Mutant analysis by rescue gene excision: New tools for mosaic studies in Drosophila.
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DOI：
10.1002/dvg.22984
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发表时间：
2016-11
期刊：
影响因子：
--
通讯作者：
Pignoni F
中科院分区：
文献类型：
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作者：
Zhou Q;Neal SJ;Pignoni F
许多经典和分子遗传工具使果蝇成为解剖基因活动的巨大模型。特别是，用于有丝分裂重组的FLP-FRT技术极大地增强了基因功能丧失的分析。该技术有效地诱导了杂合生物组织中纯合突变体克隆的形成。然而，FLP-FRT方法对细胞分裂的依赖性和其他约束条件也限制了其有效性。我们在这里描述了通过拯救基因切除（MARGE）进行突变分析的工具的生成和测试，这是一种通过在纯合突变生物体中失去拯救转基因而形成突变细胞的方法。通过使用现有的FLP、Gal4和Gal80ts试剂，可以在任何组织或细胞类型中以及在成人发育或一生中的任何时间诱导Rescue-transgene loss。同时丢失的组成荧光标记（GFP或RFP）识别突变细胞。我们证明了MARGE技术的有效性，通过翻转（克隆和圆盘宽）在图像圆盘中携带ubi - gfp的构建体，并通过在果蝇卵巢中诱导已知的yki突变表型。
A host of classical and molecular genetic tools make Drosophila a tremendous model for the dissection of gene activity. In particular, the FLP-FRT technique for mitotic recombination has greatly enhanced gene loss-of-function analysis. This technique efficiently induces formation of homozygous mutant clones in tissues of heterozygous organisms. However, the dependence of the FLP-FRT method on cell division and other constraints also impose limits on its effectiveness. We describe here the generation and testing of tools for Mutant Analysis by Rescue Gene Excision (MARGE), an approach whereby mutant cells are formed by loss of a rescue transgene in a homozygous mutant organism. Rescue-transgene loss can be induced in any tissue or cell type and at any time during development or the lifetime of the adult by using available FLP, Gal4 and Gal80ts reagents. The simultaneous loss of a constitutive fluorescence marker (GFP or RFP) identifies the mutant cells. We demonstrate the efficacy of the MARGE technique by flip-out (clonal and disc-wide) of a Ubi-GFP-carrying construct in imaginal discs, and by inducing a known yki mutant phenotype in the Drosophila ovary.