Location: Fruit and Nut ResearchTitle: Cytological and molecular characterization of three gametoclones of citrus clementina Author
Submitted to: Biomed Central (BMC) Plant Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/24/2013
Publication Date: 9/10/2013
Citation: Germana, M., Aleza, P., Carrera, E., Chen, C., Chiancone, G., Costantino, B., Dambier, D., Deng, X., Federici, C.T., Froelicher, Y., Guo, W., Ibabez, V., Juarez, J., Kwok, K., Luro, F., Machado, M., Naranjo, M.A., Navarro, L., Ollitrault, P., Rios, G., Roose, M.L., Talon, M., Xu, Q., Gmitter, F.G. 2013. Cytological and Molecular Characterization of Three Gametoclones of Citrus clementina. Biomed Central (BMC) Plant Biology. 13:129. Interpretive Summary: The sets of chromosomes of a basic number in a plant determine its ploidy level and genome complexity. A plant having two sets of similar chromosomes is called a diploid (most plants are diploid), having 3 sets called a triploid (e.g., banana and seedless watermelon), or so on. Genomes in diploids and others of higher ploidy levels can be very complex and impose challenge in genome sequencing and assembly. Haploids, only having one set of chromosomes, can occasionally be acquired by some cell culture approaches. Haploids lack the genome complexity of multiple sets of chromosomes seen in diploids and other polyploids, and are very valuable for genome sequencing and genetic studies. Identification and genetic characterization of haploid plants will greatly facilitate our understanding of their genetic integrity and selection of candidates to generate high quality reference genomes.
Technical Abstract: 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 (array-CGH). 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. 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.