Single-nuclei RNA-sequencing of plants

ABSTRACT Single-cell genomics offers a rich potential for research on plant development and environmental responses. Here, we introduce a generic procedure for plant nuclei isolation and nanowell-based library preparation for short-read sequencing. This plant-nuclei sequencing (PN-seq) method allows for analyzing the transcriptome in thousands of individual plant cells. We show the applicability of the experimental procedure to seedlings and developing flowers from Arabidopsis thaliana. The developmental trajectory of anther development is reconstructed, and stage-specific master regulators and their target genes are predicted. Novel marker genes for specific anther developmental stages are experimentally verified. The nuclei isolation procedure can be applied in different plant species, thus expanding the toolkit for plant single cell genomics experiments.