Using single-cell RNA sequencing to generate cell-type-specific split-GAL4 reagents throughout development.

Area of Science:

• Neuroscience
• Genetics
• Developmental Biology

Background:

• Accurate cell-type-specific genetic tools are crucial for understanding complex neuronal circuits and cell function.
• Existing Drosophila enhancer-based GAL4 lines often lack predictable expression patterns and specificity.
• Current split-GAL4 systems require extensive resources and time for generation and screening, with limited tools for developmental studies.

Approach:

• Leveraged single-cell RNA sequencing (scRNAseq) datasets to select gene pairs for targeted split-GAL4 line generation.
• Developed an efficient pipeline utilizing MiMIC/CRIMIC lines for generating gene-specific split-GAL4 lines via embryo injection, in vivo cassette swapping, or CRISPR knock-in.
• Validated the pipeline using the developing Drosophila visual system to demonstrate high prediction accuracy for cell-type targeting.

Key Points:

• The scRNAseq-guided pipeline efficiently generates cell-type-specific split-GAL4 lines with high accuracy.
• These novel tools enable precise labeling of cell types from early development through adulthood across various Drosophila tissues.
• The generated gene-specific split-GAL4 lines are valuable for annotating scRNAseq data and identifying novel cell types.

Conclusions:

• This work provides a powerful and efficient method for creating cell-type-specific genetic tools.
• The developed toolkit significantly advances the study of cell types in both development and adult nervous systems.
• The collection of gene-specific split-GAL4 lines represents a valuable resource for the broader fly research community.