Application of flow cytometry for genome size determination in Geosmithia fungi: A comparison of methods. Issue 10 (20th June 2014)

• Record Type: Journal Article
• Authors: Veselská, Tereza; Svoboda, Jan; Růžičková, Žaneta; Kolařík, Miroslav; Galbraith, David W.
• Is Part Of: Cytometry. Volume 85:Issue 10(2014:Oct.)
• Page Start: 854
• Page End: 861
• Publication Date: 2014-06-20
• Subjects: Flow cytometry -- Periodicals; Imaging systems in biology -- Periodicals; Imaging systems in medicine -- Periodicals; Diagnostic imaging -- Periodicals; 571.605
• DOI: 10.1002/cyto.a.22500
• Languages: English
• ISSNs: 1552-4922

Abstract

<abstract abstract-type="main"> <title>Abstract</title> <p>Genome size has played an important role in the evolution of plants and animals because changes in genome size seem to accompany if not facilitate evolutionary adaptation to environmental conditions. Flow cytometry (FCM) is a widespread method for determining genome size thanks to its high accuracy and speed of measurements. Nevertheless, only a few comparative studies of FCM methods exist in the field of mycology, and reviews are absent. In this study, we compared the suitability of several concentrations and RNAse A incubation times, fixatives and buffers for estimating genome size in fungi. We chose the genus <italic>Geosmithia</italic> as a model filamentous fungus. We also introduced a new standard, <italic>Aspergillus fumigatus</italic> CEA10, to determine absolute genome size. We found FCM to be an appropriate method for measuring genome size in fungi, but optimization steps showed that incorrect propidium iodide staining of nuclei can overestimate genome size due to cytoplasmic staining. We identified fixation with methanol:glacial acetic acid (3:1 v/v), 10% DMSO, 0.1% Triton‐X 100, and 5 mM EDTA in combination with Tris‐MgCl₂ buffer as the best treatment. © 2014 International Society for Advancement of Cytometry</p> </abstract>