Analysis of Ploidy Levels In Dichroa Febrifuga

Dichroa febrifuga Lour. is an ornamental shrub that is a member of the Hydrangeaceae.  Molecular marker data have shown that this species is closely related to Hydrangea macrophylla (Thunb.) Ser.  Fertile reciprocal hybrids between these two species have been produced, further demonstrating their close relationship.  When initial hybridizations were made, only one D. febrifuga selection (GUIZ48) was available.  Chromosome counts demonstrated that this selection was a hexaploid with 108 somatic chromosomes.  The objective of this study was to determine the ploidy level of additional D. febrifuga selections.  Nuclear DNA amounts of seven D. febrifuga selections were estimated using flow cytometry.  For each sample, at least 5000 DAPI-stained nuclei were analyzed, revealing a single peak with a coefficient of variation (CV) less than 5%.  Genome sizes were calculated as nuclear DNA content for unreduced tissue (2C) as: 2C DNA content of tissue = (mean fluorescence value of sample ÷ mean fluorescence value of standard) × 2C DNA content of standard.  Pisum sativum L. ‘Ctirad’, with a 2C content of 9.09 pg was used as the internal standard.  Flow cytometric measurements identified diploid, tetraploid and hexaploid D. febrifuga selections.  DNA content ranged from 6.1 to 7.2 pg among the three diploid selections (‘Chadwell’, ‘Yellow Wings’, an unnamed seedling from Vietnam).  Only one tetraploid selection (‘Yamaguchi Hardy’), with 12.6 pg DNA, was identified.  Three selections (GUIZ 48, ‘Yamaguchi Select’, an unnamed dwarf form) were found to be hexaploids with nuclear DNA amounts ranging from 17.0 to 17.4 pg.  Identification of diploid and tetraploid cultivars will assist in efforts to transfer desirable characteristics, such as evergreen foliage and persistent metallic blue berries, from D. febrifuga to H. macrophylla.