Abstract
Background
Three gametoclonal plants of Citrus clementina Hort. ex Tan., cv. Nules, designated ESP,
FRA, and ITA (derived from three labs in Spain, France, and Italy, respectively), were
selected for cytological and molecular characterization in order to elucidate genomic
rearrangements provoked by haploidization. The study included comparisons of their ploidy,
homozygosity, genome integrity, and gene dosage, using chromosome counting, flow
cytometry, SSR marker genotyping, and array-Comparative Genomic Hybridization (arrayCGH).
Results
Chromosome counting and flow cytometry revealed that ESP and FRA were haploid, but ITA
was tri-haploid. Homozygous patterns, represented by a single peak (allele), were observed
among the three plants at almost all SSR loci distributed across the entire diploid donor
genome. Those few loci with extra peaks visualized as output from automated sequencing
runs, generally low or ambiguous, might result from amplicons of paralogous members at the
locus, non-specific sites, or unexpected recombinant alleles. No new alleles were found,
suggesting the genomes remained stable and intact during gametogenesis and regeneration.
The integrity of the haploid genome also was supported by array-CGH studies, in which
genomic profiles were comparable to the diploid control.
Conclusions
The presence of few gene hybridization abnormalities, corroborated by gene dosage
measurements, were hypothetically due to the segregation of hemizygous alleles and minor
genomic rearrangements occurring during the haploidization procedure. In conclusion, these
plants that are valuable genetic and breeding materials contain completely homozygous and
essentially intact genomes
