Targeted cybridization in citrus: transfer of Satsuma cytoplasm to seedy cultivars for potential seedlessness

Abstract

CMS (cytoplasmic male sterility) can be controlled by the mitochondrion genome in higher plants, including Satsuma mandarin. Somatic fusion experiments in citrus combining embryogenic callus protoplasts of one parent with leaf protoplasts of a second parent often produce cybrid plants of the leaf parent, a phenomenon occurring most often with interspecific fusion combinations. In an attempt to practically exploit this cybridization phenomenon, we conducted somatic fusion experiments combining embryogenic suspension-derived protoplasts of Satsuma mandarin, Citrus unshiu Marc. cv. Guoqing No. 1 (G1), a male-sterile cultivar, with leaf protoplasts of other seedy types--Hirado Buntan Pink pummelo (HBP) [Citrus grandis (L.) Osbeck], Sunburst mandarin (C. reticulata Blanco), Orie Lee hybrid (C. reticulata cv. Clementine x Murcott tangor), and Murcott tangor [C. reticulata x C. sinensis (L.) Osbeck], respectively--in an attempt to generate seedless cybrids by the targeted transfer of CMS. The genetic identities of regenerated plants from all four parental combinations were determined by flow cytometry, SSR, CAPS (or PCR-RFLP), RFLP, and chloroplast-SSR analyses. Regenerated plants from the first three parental combinations were diploids, and the cybrid nature of G1 + HBP with the mitochondrion genome from G1 and the chloroplast genome from HBP was confirmed, whereas the cybrid nature of the remaining two combinations was difficult to confirm because of the close phylogenetic relatedness of both fusion parents, as expected. Plants from G1 + Murcott were confirmed as tetraploid somatic hybrids. This is the first report of targeted citrus cybrid production by symmetric fusion with male-sterile Satsuma as the callus parent and other seedy cultivars as the leaf parents.

Segregation and Heritability of Male Sterility in Populations Derived from Progeny of Satsuma Mandarin

“…Recent studies focused to the involvement of CMS for the male sterility of Satsuma [12–16,28]. However, the observed segregation and higher heritability for the male sterility in this study strongly suggest the indispensable role of nuclear gene to this important trait.…”

“…These studies led following studies that the presumed male sterility of Satsuma would be achieved by CMS. Guo and colleagues had reported efforts to introduce the male sterility of Satsuma to other varieties by transferring cytoplasm of it by cybrid formation [12–15]. Though these studies did not confirm male sterility in these cybrid plants, recent studies by Zhen et al reported severe retardation of anther was observed in the cybrid pomelo, and proposed interaction of dysfunctional mitochondrial genes to suppress nuclear genes to suppress anther formation [16].…”

Comparative Transcript Profiling of a Male Sterile Cybrid Pummelo and Its Fertile Type Revealed Altered Gene Expression Related to Flower Development

“…Considering the possibility of targeted mitochondrial genome transfer and the importance of seedless trait for fresh citrus market, we put forward a strategy to transfer male sterile and seedless trait by somatic cybridization in efforts to reduce the seed content of important cultivars [9]. In our previous study, diploid cybrids were generated [10] by transferring the male sterile cytoplasm of Satsuma mandarin ( Citrus unshiu Marc. ), a seedless variety with CMS character [11], to seedy citrus cultivars.…”

Comparative transcript profiling of gene expression between seedless Ponkan mandarin and its seedy wild type during floral organ development by suppression subtractive hybridization and cDNA microarray

“…somatic hybridization) have potential to effectively expedite breeding process of citrus [13-15]. As most citrus varieties can produce fruits parthenocarpically [16], male or female sterility, embryo sac abortion, self-incompatibility, polyploidy and even environmental stress can result in seedless citrus fruits [17,18]. Actually there were some successful reports about seedless fruit production by genetic transformation.…”