Mastering DNA Content Estimation by Flow Cytometry as an Efficient Tool for Plant Breeding and Biodiversity Research

Abstract

Flow cytometry gives a unique opportunity to analyze thousands of individual cells for multiple parameters in a course of minutes. The most commonly used flow cytometry application in plant biology is estimation of nuclear DNA content. This becomes an indispensable tool in different areas of plant research, including breeding, taxonomy, plant development, evolutionary biology, populational studies and others. DNA content analysis can provide an insight into natural ploidy changes that reflect evolutionary processes, such as interspecific hybridization and polyploidization. It is also widely used for processing samples with biotechnologically induced ploidy changes, for instance, plants produced by doubled haploid technology. Absolute genome size data produced by cytometric analysis serve as useful taxon-specific markers since genome size vary between different taxa. It often allows the distinguishing of species within a genus or even different subspecies. Introducing flow cytometry method in the lab is extremely appealing, but new users face a significant challenge of learning instrument management, quality sample preparation and data processing. Not only is flow cytometry a complex method, but plant samples have unique features that make plants a demanding research subject. Without proper training, researchers risk damaging the expensive instrument or publishing poor quality data, artifacts or unreproducible results. We bring together information from our experience, key papers and online resources to provide step by step protocols and give a starting point for exploring the abundant cytometry literature.