Protocol to produce and apply barcoded rabies virus for single-neuron input mapping in mice

Area of Science:

• Neuroscience
• Molecular Biology
• Genomics

Background:

• Understanding neural circuits is crucial for neuroscience.
• Current methods for mapping neuronal inputs have limitations in throughput and integration with molecular data.
• Rabies virus tracing is a powerful tool for studying neuronal connectivity.

Purpose of the Study:

• To present a detailed protocol for producing and applying barcoded rabies virus for single-neuron input mapping in mice.
• To enable high-throughput mapping of brain-wide inputs to individual neurons.
• To facilitate integration of neural tracing with transcriptomic profiling.

Main Methods:

• Generation of barcode plasmid libraries.
• Production of barcoded rabies virus.
• Isolation of target brain regions.
• Preparation of single-cell suspensions or bulk RNA.
• Construction of barcode amplicon libraries from input regions.
• Initial analysis of sequencing data.

Main Results:

• The protocol enables high-throughput mapping of brain-wide inputs to individual neurons.
• The method allows for integration with transcriptomic profiling.
• Detailed steps for virus production, tissue processing, and data analysis are provided.

Conclusions:

• Barcoded rabies viral tracing is an effective method for comprehensive neural circuit analysis.
• This protocol provides a valuable tool for researchers studying brain connectivity and gene expression.
• The technique advances the field of neuroscience by enabling detailed single-neuron connectomics.