Triple-color FISH chromosome constitution and genome size analyses confirmed the ploidy of newly developed ten Hibiscus sp. cultivars with different breeding history

Hibiscus syriacus L. (Rose of Sharon), is the national flower of Korea. H. syriacus has both ornamental and medicinal value, and many cultivars have been exploited for its development. Cytogenomic information is crucial for elucidating chromosomal composition, ploidy, and specific sequence distribution. In this study, chromosome constitution, genome size, and ploidy were analyzed in ten Hibiscus sp. cultivars with different breeding histories using triple-color fluorescence in situ hybridization (FISH) and flow cytometry. Prelabeled oligonucleotide probes (PLOPs) of three universal tandem repeats (5S rDNA, 45S rDNA, and telomeric DNA) were used for FISH. The chromosome length ranged from 1.20 to 4.84 μm, and the chromosome number varied from 2n = 84 to 2n = 172 with the basic chromosome number x = 21. Based on chromosome number, FISH signals, and genome size, three cultivars were identified as euploids of tetraploid and hexaploid euploids, and the remaining seven cultivars as aneuploids of tetraploid, hexaploid, and octoploid euploids. The number of 5S rDNA signals was two, three, and four for tetraploid, hexaploid, and octoploid, respectively. Whereas 45S rDNA signals varied from two to five pairs among the cultivars. Genome size (2C) ranged from 4.35 to 4.80 pg in tetraploids, 6.35 to 6.78 pg in hexaploids, and 8.20 to 8.94 pg in octoploids, which corresponded to the ploidy identification using FISH chromosome composition. FISH using PLOPs demonstrated that the nuclear DNA content increased proportionally with the ploidy level, indicating that the chromosome composition and genome size were consistent. The findings will be useful for future genomic research on Hibiscus species breeding and development strategies.