Self-(in)compatibility in Tunisian apple accessions [Malus domestica. Borkh]: S-genotypes identification and pollen tube growth analysis.

MAIN CONCLUSION: Self-incompatibility studies have revealed a potential use of Tunisian apple resources for crop improvement and modern breeding programs and a likely correlation between the pollen tube growth and flowering period. Apples [Malus domestica. Borkh] exhibit an S-RNase-based gametophytic self-incompatibility (GSI) system. Four primer combinations were used to S-genotype eighteen Tunisian local apple accessions and twelve introduced accessions that served as references. Within the Tunisian local accessions, S2, S3, S7, and S28 S-alleles were the most frequent and were assigned to 14 S-genotypes; among them, S7S28, S3S7, S2S5, and S2S3 were the most abundant. PCA plot showed that population structuring was affected by the S-alleles frequencies and revealed a modern origin of the Tunisian varieties rather than being ancient ones. Nonetheless, the results obtained with 17 SSR markers showed a separate grouping of local Tunisian accessions that calls into question the hypothesis discussed. Pollination experiments showed that the pollen started to germinate within 24 h of pollination but 48 h after pollination in the "El Fessi" accession. The first pollen tubes arrived in the styles within 36 h of pollination in two early flowering accessions known as "Arbi" and "Bokri", and after 72 h of pollination in late flowering "El Fessi" and 48 h after pollination in remaining accessions. The first pollen tube arrests were observed in accessions "Arbi" and "Bokri" within 84 h of pollination, within 108 h of pollination in "El Fessi" and within 108 h of pollination in remaining accessions. In the apple accession called "Boutabgaya," the pollen tubes reached the base of the style within 120 h of pollination without being aborted. Nevertheless, the self-compatible nature of "Boutabgaya" needs more studies to be confirmed. However, our results revealed the malfunction of the female component of the GSI in this accession. To conclude, this work paved the path for further studies to enhance the insight (i) into the relation between the flowering period and the pollen tube growth, (ii) self-compatible nature of "Boutabgaya", and (iii) the origin of the Tunisian apple.

Molecular and Evolutionary Characterization of Pollen S Determinant (SFB Alleles) in Four Diploid and Hexaploid Plum Species (Prunus spp.).

In more than 60 families of angiosperms, the self- and cross-fertilization is avoided through a complex widespread genetic system called self-incompatibility (SI). One of the major puzzling issues concerning the SI is the evolution of this system in species with complex polyploid genomes. Among plums, one of the first fruits species to attract human interest, polyploid species represent enormous genetic potential, which can be exploited in breeding programs. However, molecular studies in these species are very scarce due to the complexity of their genome. In order to study the SFB gene [the male component of gametophytic self-incompatibility system (GSI)] in plum species, 36 plum accessions belonging to diploid and hexaploid species were used. A total of 19 different alleles were identified; 1 of them was revealed after analyzing sequences. Peptide sequence analysis allowed identifying the five domains features of the SFB gene. Polymorphism analysis showed a subtle difference between domesticated and open pollinated Tunisian accessions and suggested a probable influence of the ploidy level. Divergence analysis between studied sequences showed that a new specificity may appear after 5.3% of divergence at synonymous sites between pairs of sequences in Prunus insititia, 6% in Prunus cerasifera, 8% and 9% in Prunus domestica and Prunus salicina respectively. Furthermore, sites under positive selection, the ones more likely to be responsible for specificity determination, were identified. A positive and significant Pearson correlation was found between the divergence between sequences, divergence time, fixed substitutions (MK test), and PSS number. These results supported the model assuming that functionally distinct proteins have arisen not as a result of chance fixation of neutral variants, but rather as a result of positive Darwinian selection. Further, the role that plays recombination can not be ruled out, since a rate of 0.08 recombination event per polymorphic sites was identified.

Analysis of Self-Incompatibility and Genetic Diversity in Diploid and Hexaploid Plum Genotypes.

During the last decade, S-genotyping has been extensively investigated in fruit tree crops such as those belonging to the Prunus genus, including plums. In plums, S-allele typing has been largely studied in diploid species but works are scarcer in polyploid species due to the complexity of the polyploid genome. This study was conducted in order to analyze the S-genotypes of 30 diploid P. salicina, 17 of them reported here for the first time, and 29 hexaploid plums (24 of P. domestica and 5 of P. insititia). PCR analysis allowed identifying nine S-alleles in the P. salicina samples allocating the 30 accessions in 16 incompatibility groups, two of them identified here for the first time. In addition, pollen tube growth was studied in self-pollinated flowers of 17 Tunisian P. salicina under the microscope. In 16 samples, including one carrying the Se allele, which has been correlated with self-compatibility, the pollen tubes were arrested in the style. Only in one cultivar ("Bedri"), the pollen tubes reached the base of the style. Twelve S-alleles were identified in the 24 P. domestica and 5 P. insititia accessions, assigning accessions in 16 S-genotypes. S-genotyping results were combined with nine SSR loci to analyze genetic diversity. Results showed a close genetic relationship between P. domestica and P. salicina and between P. domestica and P. insititia corroborating that S-locus genotyping is suitable for molecular fingerprinting in diploid and polyploid Prunus species.

Combination of Simple Sequence Repeat, S-Locus Polymorphism and Phenotypic Data for Identification of Tunisian Plum Species (Prunus spp.).

Plums (Prunus spp.) are among the first fruit tree species that attracted human interest. Artificial crosses between wild and domesticated species of plums are still paving the way for creation of new phenotypic variability. In Tunisia, despite a considerable varietal richness of plum as well as a high economic value, the plum sector is experiencing a significant regression. The main reason of this regression is the absence of a national program of plum conservation. Hence, this work was aimed to phenotypically and genetically characterize 23 Tunisian plum accessions to preserve this patrimony. Closely related Prunus species from the same subgenus may be differing at two characteristics: ploidy level and phenotypic traits. In this study, single sequence repeat (SSR) markers allowed distinguishing between eighteen diploid accessions and five polyploid accessions, but SSR data alone precluded unambiguous ploidy estimation due to homozygosity. In contrast, S-allele markers were useful to identify the ploidy level between polyploid species, but they did not distinguish species with the same ploidy level. Seven out of 12 phenotypic traits were shown to be discriminant traits for plum species identification. Molecular and phenotypic traits were significantly correlated and revealed a powerful tool to draw taxonomic and genotypic keys. The results obtained in this work are of great importance for local Tunisian plum germplasm management.