Single-Cell RNA-Sequencing in Nodulated Roots of Medicago truncatula.

This section details the optimized methods for single-cell RNA sequencing (scRNA-seq) in Medicago truncatula roots and nodules. We describe a detailed experimental workflow, comprising plant growth and bacteria application, protoplast isolation, quality control, and library processing according to 10× Genomics®, suitable for both long- and short-read sequencing. This protocol can serve as a starting point for establishing robust, high-quality scRNA-seq workflows to study nodulation and root development in legumes.