Best practices in plant cytometry

Abstract

Best practices in plant cytometry Flow cytometry (FCM) and flow cytometric sorting (FCS) systems have developed as experimental tools of remarkable power and are enjoying an ever-increasing impact in the general field of biology. 1 Application of these tools to plant biology has developed more slowly given that the natural form of plants infrequently resembles that of the single cell suspension, prototypically the hematopoietic system that drove the original development of FCM/FCS. Nevertheless, these systems have had a profound influence at all levels of plant biology, from the study of single cells and subcellular organelles, to the behavior of populations of plants, and ultimately to the performance of ecosystems. It is safe to say their impact has not plateaued, as further applications of this unique technology are increasingly developed by innovative scientists around the world to address questions both in the basic sciences, and to increasingly confront emerging problems in the applied sector. For example, in addressing the challenges of sustainable production of sufficient food resources based on plant breeding involving ploidy-based approaches (e.g., induction of polyploidy) 2 for the needs of our future global citizens, FCM, and FCS systems will play central roles in this effort. The degree to which FCM and FCS systems have impacted plant biology and applied agricultural sciences must not be understated. The major applications of DNA FCM are ploidy level and genome size estimations, and cell cycle analysis/endoreplication (with the later included in a lower percentage of studies). Indeed, FCM is currently/ extensively and almost exclusively employed as the method of choice for measurement of plant genome sizes. 3,4 Measurements of this type impact agriculture in terms of ploidy estimation, with applications ranging from plant biotechnology, breeding and seed quality testing to taxonomy and population biology. They also impact the fundamental plant sciences in terms of biosystematics, ecology, evolution, genomics, and conservation, among other applications. One of the most startling observations of the angiosperms is the bandwidth occupied by genome size, which spans almost 2400-fold. Flow sorting of higher plant chromosomes has provided invaluable information regarding the organization of DNA sequences within plant species. It has also greatly facilitated the process of wholegenome sequencing by permitting subdivision of large genomes into samples comprising entire chromosomes or chromosome arms. 5 FCS methods applied to wall-less cells (protoplasts) expressing fluorescent proteins (FPs) in a cell type-specific manner have allowed elucidation of patterns of co-regulated gene expression and plant hormone gradients identification 6,7 within organized tissues, such as roots. 8,9

Synergistic Action of Mild Heat and Essential Oil Treatments on Culturability and Viability of Escherichia coli ATCC 25922 Tested In Vitro and in Fruit Juice

“…The study highlighted the importance of combining traditional culture-based analytical methods with fast and high-throughput ones for monitoring the microbiological safety of fruit juice. The comparison (solely on an economic basis) between the classic microbiological technique and flow cytometry for food applications clearly works in favor of the classic technique, but FCM is a tool which extends microbiological analyses capabilities and has a huge number of applications in several fields [ 65 , 66 , 67 ], and diminishing costs and user-friendly software are increasingly promoting its use. Flow cytometry proved to be capable of providing information on a large number of cells during the different phases of their growth, exceeding the limitation of the traditional culture-based methods, which underestimated bacterial growth.…”

Validation of crowd‐sourced plant genome size measurements

“…An obvious issue is the uncovering ex post facto of critical variables that had not been recorded in the databases and that now are no longer available. For further discussions of Best Practices in Plant Cytometry, please visit Galbraith et al [13], and references cited therein.…”

“…For further discussions of Best Practices in Plant Cytometry, please visit Galbraith et al [13], and references cited therein.…”

Diversity and Cytogenomic Characterization of Wild Carrots in the Macaronesian Islands

“…A comprehensive understanding of plant genomes is still largely missing, due to the relatively small number of species examined to date (around 12,273 species) [ 1 ]. High-throughput flow cytometry (FCM) screens for DNA content have especially aided in identifying the frequency of polyploids [ 2 ], as well as their phenotypic and geographical distributions [ 3 ]. Advances in flow cytometry are being applied to study intraspecific and interspecific variation (e.g., [ 4 ]), as well as to resolve complex low-level taxonomies, including the delimitation of species boundaries [ 5 ].…”