An Optimised Translating Ribosome Affinity Purification (TRAP) Protocol for Low-Abundance Drosophila tissues

This is especially important in cells with extremely elaborated architectures such as neurons. We present a highly sensitive TRAP protocol optimised for the specific isolation of ribosome-bound mRNAs in Drosophila from limited input samples, which enabled the isolation of axonal mRNAs from larval and adult (leg) motor neurons.

RNA-seq revealed a set of axonally translated transcripts, including mRNAs encoding multiple ribosomal and mitochondrial proteins. Notably, these are among the types of transcripts found in axons of other species, indicating that axonal translation is conserved in Drosophila. Our optimised low-input TRAP method will allow the study of local translation to be explored in conjunction with Drosophila genetic manipulations, facilitating in-depth investigation of axonal translation across genetic backgrounds, developmental stages, and experimental conditions such as models of human disease.