Protocol for replacing coding intronic MiMIC and CRIMIC lines with T2A-split-GAL4 lines in Drosophila using genetic crosses

Abstract

Summary: Here, we present a protocol for generating gene-specific split-GAL4 drivers from coding intronic Minos-mediated integration cassette/CRISPR-mediated integration cassette (MiMIC/CRIMIC) lines in Drosophila. We describe steps for four rounds of in vivo genetic crosses, PCR genotyping, and fluorescence imaging to ensure correct orientation of split-GAL4 integration before establishing stable fly stocks. This protocol offers a cost-effective alternative to traditional microinjection techniques for converting coding intronic MiMIC/CRIMIC lines into gene-specific split-GAL4 lines that are adaptable for fly researchers working on different tissues.For complete details on the use and execution of this protocol, please refer to Chen et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Developmental biology
• Genetics
• Model Organisms
• Neuroscience