Single-Cell and Spatial Transcriptomic Analysis of Maize Embryo Development: a Sample Preparation Protocol.

Abstract

Maize is an important crop that contributes to the modern economy in various ways, including use for human consumption, as animal feed, and in industrial products. Research on maize is crucial for understanding plant development, which in turn provides valuable insight into improvement of maize crops to meet the food demands of a growing population. Maize embryogenesis, which is the primordial stage of the corn life cycle, determines the fundamental body plan and developmental programs that organize the tissue patterning and subsequent growth and reproduction of the corn plant. Investigating maize embryogenesis at high cellular resolution can enhance our understanding of the homology, ontogeny, and developmental genetic mechanisms of embryonic organ morphogenesis. However, until recently, no published studies have used methods for analyzing maize embryo development at single-cell resolution. This protocol describes single-cell RNA sequencing (scRNA-seq) and spatial transcriptomic analyses, which are powerful, combinatorial tools that can be used to study maize embryogenesis at the single-cell level within a spatial context. These tools have the power to reveal transcriptomic relationships between tissues/organs, and to provide insight into the gene regulatory networks operating during embryogenesis. In this protocol, we describe a detailed procedure to prepare maize embryo samples for construction of scRNA-seq and Visium spatial transcriptomic libraries that are suitable for massively parallel sequencing. Our protocol borrows from prior published studies and manufacturer's instructions and is optimized for studies of the maize embryo.