RRC ID 79032
Author Phipps DN, Powell AM, Ables ET.
Title Utilizing the FLP-Out System for Clonal RNAi Analysis in the Adult Drosophila Ovary.
Journal Methods Mol Biol
Abstract The ability to conduct spatially controlled RNA interference (RNAi) for gene knockdown using the UAS/Gal4 system is among the most appealing techniques available for analysis of gene function in the Drosophila ovary. While gene knockdown experiments in somatic cells in the developing organism (i.e., embryos and larvae) are effectively and commonly performed, the use of RNAi in adult ovarian cells can be hampered by the unintended deleterious effects of Gal4 expression in "off-target" developing tissues. Mosaic analysis overcomes these problems by imparting temporal and spatial control over gene manipulation, providing a useful tool to compare manipulated cells with wild-type cells in the same tissue. Here, we provide a method to utilize the UAS/Gal4 system in combination with the Flippase (FLP)-Flippase Recognition Target (FRT) system to generate positively labeled "FLP-Out" clones expressing a chosen RNAi in both the germline and the soma in the Drosophila ovary. This protocol outlines each step of the generation of clones and the selection of appropriate fly stocks and reagents, providing a guide to this powerful tool in the Drosophila genetic toolbox. These techniques allow for RNAi analysis within a specific cell type, providing an opportunity to study a variety of unique aspects of cell function that would not be possible in more traditional RNAi-based experiments.
Volume 2626
Pages 69-87
Published 2023-1-1
DOI 10.1007/978-1-0716-2970-3_4
PMID 36715900
PMC PMC10044525
MeSH Animals Drosophila* / genetics Drosophila* / metabolism Drosophila Proteins* / genetics Drosophila Proteins* / metabolism Drosophila melanogaster / genetics Female Genetic Techniques Ovary / metabolism RNA Interference