Estimation of chromosome number among the progeny of a self-pollinated population of triploid Senno (Lychnis senno Siebold et Zucc.) by flow cytometry

Abstract

Flow cytometry is a powerful tool to determine ploidy level and is commonly applied to measure DNA content and analyze the cell cycle. Here, we analyzed ploidy among the progeny derived from a population of self-pollinated triploid Senno (Lychnis senno Siebold et Zucc.). Each plant within the progeny was estimated to be an aneuploid showing a different ploidy level. For a large population, identification of chromosome number by chromosome observation is time-consuming. Therefore, a simple procedure to categorize different ploidy levels and chromosome number is required to facilitate breeding efforts. We applied flow cytometry to estimate chromosome number using internal standards for reference; these estimates were compared with the results obtained following chromosome observation. Leaf samples from progeny of triploid Senno were measured by flow cytometry in combination with an internal standard to evaluate DNA content accurately. By using appropriate internal standards, chromosome number was effectively estimated. When the parent triploid plant (strain MS, 2n = 3x = 36) was used as the internal standard, diploid (2n = 24) and different aneuploids (2n = 25, 44, 45, 55) were discriminated. For measurement of presumed triploids, strain No. 27 (2n = 25) was used as another internal standard. As a result, triploid (2n = 3x = 36) and different aneuploids (2n = 33, 35) were detected. The exact number of chromosomes as per chromosome observation corresponded well with the number estimated through flow cytometry. This procedure will facilitate ready, accurate, and reliable estimation of chromosome number using an adequate reference sample for internal standard. We discuss the possibility to estimate chromosome number by flow cytometry and the applicability of the procedure for determining the ploidy level as well as aneuploids in large populations derived from parental triploid Senno plants.