Analysis of genetic diversity among some Persian walnut genotypes (Juglans regia L.) using morphological traits and SSRs markers

There is a high diversity among cultivated walnut trees in Iran due to its long time of seed propagation and vast area of cultivation. In this study some morphological characters as well as Simple Sequence Repeat (SSRs) markers were used to analyze the genetic diversity and relationships among 31 Iranian walnut genotypes along with four foreign cultivars. The nut weight ranged from 7.52 to 17.73 g, kernel weight from 4.00 to 9.83 g, and kernel percentage ratio from 38.78 to 67.05% among studied genotypes. In SSRs analysis, nine primer pairs were tested that produced 39 alleles ranging from 2 to 8, with a mean value of 5.10 allele per primer. The Iranian genotypes showed relatively high diversity both for their SSRs loci and morphological traits. Although the foreign cultivars (‘Serr’, ‘Vina’, ‘Franquette’ and ‘Lara’) clustered with each other, they also laid close and within the Iranian genotype. The results of the study provided us with valuable diversity among our genotypes which could be used for breeding studies and also showed the power of genetic markers for analysis and evaluation of this diversity.     

Physical and chemical characterization of Ceratonia siliqua L. germplasm in Turkey

Carob (Ceratonia siliqua L.) is natively grown in the coastal areas of the Aegean and Mediterranean region in Turkey. The main carob growing areas (Mediterranean and Aegean) were surveyed and a total 70 promising wild and grafted carob genotypes were selected based on physical and chemical pod properties. Preliminary results showed that Turkey has very rich carob genetic resources containing considerable variations for most of the pod traits. In general, wild carob had smaller pods than the grafted ones. Pod mass ranged from 29.16 g to 120.28 g in the wild genotypes and 71.71 g to 147.91 g in the grafted genotypes with an average 107.61 g for the grafted and 63.72 g for the wild genotypes. The average pod dimensions (width, length and thickness) were 18.16 mm, 16.18 cm and 6.42 mm for the wild genotypes and 21.79 mm, 16.91 cm and 8.79 mm for the grafted genotypes. The average soluble solid content ranged from 59.42% in the wild and 64.06% in the grafted genotypes. The average total acidity of the wild and grafted genotypes was similar (0.78%). The pod traits of the wild genotypes differ significantly from those of the grafted genotypes, and the wild genotypes showed higher variations in these traits compared to the grafted genotypes.     

Bacterial canker of olives caused by of Pseudomonas syringae pv. syringae in Iran

Guilan is one of the main province olive oil producers in Iran. The culture of olive trees is of considerable importance. One problem that affects Iran olive orchards is a disease bacterial canker caused by Pseudomonas syringae pv. syringae. In this research, the bacterium was recovered from sunken brown stem lesions on 2-year-old olive trees cv. Marie, during survey from olive orchards in different areas of Guilan province. Samples were taken from infected tissues and were washed with sterile distilled water and crushed in peptone water. Then 50 μl of the extract was cultured on NA and King's B medium containing cyclohexamid antibiotic (50 μg/ml). After 48–72 h, bacterial colonies were selected. Based on morphological, physiological, biochemical, pathogenicity properties and PCR methods with specific primers the predominant pathogenic type was identified as P. s. pv. syringae. This is the first report of the existence of P. s. pv. syringae on olive trees in Iran.     

Comparative analysis of genetic diversity in Prunus L. as revealed by RAPD and SSR markers

RAPD and SSR markers were used for genetic diversity evaluations among 15 genotypes selected from the genus Prunus L. Altogether 40 RAPD primers and 21 primer pairs designated for microsatellite loci were applied on the whole group of genotypes.     

Molecular polymorphism of cytoplasmic DNA in Ficus carica L.: Insights from non-coding regions of chloroplast DNA

The trnL (UAA) intron and the intergenic spacer between the 3′ exon of trnL (UAA) and trnF (GAA) sequences were used as genetic markers for differentiating Ficus carica cultivars and establishing refined genetic relationships. The study was based on 20 fig cultivars, collected from south and centre of Tunisia. Since, the intron was thought to be more variable among close relatives than is the chloroplast spacer. The size of these non-coding regions varied from 554 to 589 and from 989 to 1022 bases pairs for the intron and the combined sequences correspondingly. The average of GC content was 33.9% and 34.6% in the intron and the combined intron and spacer respectively. High values of A + T contents were detected in both data sets and may explain the high proportions of transversions founded. The observed variation pattern of plastid DNA provides evidence of an important genetic diversity. The overall transition/transversion bias (R) was 0.202 in the intron and 0.27 in the combined regions. The RI index of 0.592 indicates that these combined sequences have clearly more homoplasy then the intron (RI = 0.705) and spacer (RI = 0.777) sequences separately. Phylogenetic trees were generated based on maximum parsimony (MP) and neighbor-joining (NJ) analysis of the chloroplast sequences data. Results proved that a typically continuous genetic diversity characterizes the local fig germplasm. In fact, relationships inferred from the cpDNA analysis suggest several clades, which do not show geographical or tree sex correspondence. Although the level of apparent diversity is considerable, we may conclude that non-coding regions of chloroplast genome provide a new and practical opportunity to evaluate genetic diversity and to discriminate fig cultivars. Revealed cytoplasmic DNA markers are reliable to elaborate a molecular data base to conduct management and breeding programs on local fig germplasm.     

Comparative analysis of genetic diversity in Tunisian apricot germplasm using AFLP and SSR markers

Eighty-one accessions representing apricot germplasm in Tunisia were collected from different areas of cultivation and fingerprinted using amplified fragment length polymorphism (AFLP) and microsatellites (SSR) markers. A total of 339 polymorphic markers were revealed using 5 AFLP primers combinations and 24 SSR loci. AFLP and SSR markers expressed a high level of polymorphism allowing the distinction of the accessions with an efficiency coefficient of discrimination of 100% for AFLP and 97% for SSR markers. Genetic diversity structure was assessed with AFLPs and SSRs markers separately then with combined matrix data by the help of hierarchical clustering elaborated using Wards method based on Nei and Li (1979) distances. Comparison of the obtained dendrograms revealed a phylogeographic structure into two major groups with significant conservation between the observed subgroups in relation with the geographic origin of the accessions. The relative efficiency of the markers in determining the genetic relationships among apricot accessions has been assessed and a combination of AFLPs and SSRs markers was the most effective. In addition, Mantel test based on genetic distances indicated highly significant correlation between AFLP-SSR data and each of the AFLP and SSR ones, with Pearson correlation values of r = 0.873 and r = 0.692, respectively, revealing the higher efficiency of the combination of both molecular techniques (AFLP and SSR) to estimate the levels of genetic variability among apricot germplasm.     

Germplasm diversity and genetic relationships among walnut (Juglans regia L.) cultivars and Greek local selections revealed by Inter-Simple Sequence Repeat (ISSR) markers

Inter-Simple Sequence Repeat (ISSR) markers were used for the assessment of genetic diversity among walnut (Juglans regia L.) selections from Greek native populations in comparison to internationally cultivated walnut genotypes. Similarity coefficient values from 0.13 to 0.93 (with an average of 0.48) were found among the 56 accessions examined, which indicated the presence of a high degree of genetic variability. Most international cultivars were grouped together while most Greek native populations could not be placed into a distinct group. The Greek native population genotypes were found more diverse than the international cultivars. The mean similarity coefficient values for the former and latter were 0.44 and 0.56, respectively. In the cultivar group, two subgroups were distinguished; one consisted of genotypes involving ‘Payne’ and the other ‘Franquette’ in their pedigrees. Some cultivars and populations could not be grouped according to their pedigrees or collection area. Analysis of molecular variance (AMOVA) revealed that a larger part of the genetic variation exists among Greek walnut populations within a collection region (89%) than among the regions (11%). The pairwise regional PhiPT values indicated that the most geographically distant regions are the most genetically differentiated. The high variability existing in the Greek germplasm in combination with their valuable agro-morphological traits suggested that it would be beneficial to utilize this native germplasm pool in walnut breeding programs and germplasm management activities to maximize genetic diversity in cultivated walnut.     

Phenotypic variations in fruits and selection potential in Sclerocarya birrea subsp. birrea

The domestication of indigenous fruits through agroforestry is seen as one of the important issues in the transformation in the way in which land is used within rural areas. A study of the variation in fruit traits is also important as it is a prerequisite for cultivar development in the domestication process. Following an aridity gradient, phenotypic variation was assessed in fruits of Sclerocarya birrea subsp. birrea, native to West African semi-arid areas. Fruits were collected from 42 trees of various diameters in agroforestry parklands from the dry and semi-humid Sudanian zones. They were partitioned into peel, flesh and pit. Each fruit was labelled and its components were measured and weighted. The overall mean fruit mass was 18.58 ± 0.24 g (mean ± SE) but fruits from the population in the drier zone were significantly larger (19.90 ± 0.37 g vs 17.02 ± 0.24 g; P < 0.001). The results showed a strong correlation between fruits and their components (P < 0.05). Tree diameter was very weakly correlated with fruit traits. There was a high level of variation in fruit characteristics and components within and between populations. The within-population variation accounted for the greatest part (67–100%) of the total variation. Many trees, mainly from the drier zones, showed superior phenotypic traits. Five groups of trees which represented different fruit morphotypes were identified for various prospective exploitations. The results strongly support the implementation of preliminary practical conservation action and domestication of S. birrea in West Africa.     

Use of ISSR,SRAP, and RAPD markers to assess genetic diversity in Turkish melons

The genetic relationships among 63 melon (Cucumis melo L.) genotypes collected from various regions of Turkey were determined by comparing their molecular ISSR, SRAP, and RAPD markers with those of 19 foreign melon genotypes to investigate the taxonomic relationships and genetic variation of Turkish melon germplasm. Total 162 polymorphic markers (69, 18, and 75 obtained from ISSR, SRAP, and RAPD primers, respectively) were used to define the genetic similarity among the melon genotypes by dendrogram or two and three dimensional scalings. The average similarity (SM coefficient) between any two pairs of accessions examined as estimated by molecular variation was 0.73 ± 0.48. Within-group genetic similarities ranged between 0.46 and 0.96. Related genotypes or genotypes collected from similar regions were partitioned to similar clusters. Southeastern Anatolian genotypes were distinctly apart from group inodorus and group cantalupensis (sweet) genotypes. This reinforced the position of Turkey in the secondary genetic diversity center of melon. The genetic diversity among Turkish genotypes (H = 0.28 and I = 0.42) was only a little less than that of the world accessions (H = 0.30 and I = 0.45). On the other hand, the percentage of polymorphic loci among Turkish melon genotypes (90.7%) was even higher than that of the world accessions (87.6%).     

Comparison of the use of morphological,protein and DNA markers in the genetic characterization of Iranian wild Prunus species

In this study, the genetic diversity of four Iranian wild Prunus species including Prunus eleagnifolia, Prunus hauskenchtii, Prunus scoparia and Prunus lycioides were investigated using morphological, protein and DNA markers. DNA markers included nuclear and chloroplast SSRs and self-incompatibility (S) allele amplification. At the morphological level, leaf width showed significant differences between the four wild Prunus species. Concerning fruit and kernel characters, their values are relatively similar indicating the high degree of homoplasy described in Prunus. Nuclear SSR markers have been the most abundant markers with a higher polymorphism in comparison with morphological, protein and chloroplast SSR markers. Results also indicated the high variability present in the S locus. On the other hand, the correlation between the clustering based on DNA markers and protein were in general low. Dendogram performed using nuclear and chloroplast SSR indicated a more diffuse clustering between the wild almond species probably due to the natural introgression of genes observed in these wild almond species. Data from the analysis of the total protein seems to be more accurate to establish taxonomy relationships in these very close wild species.     

Effects of water deficit on the vegetative response,yield and oil quality of olive trees (Olea europaea L., cv Coratina) grown under intensive cultivation

An experiment was carried out in a young high-density olive grove (556 plants ha⁻¹—Olea europaea L., cv Coratina) located in Southern Italy to evaluate the effect of different soil water availability on the vegetative and productive performances of olive trees also looking into the quality of the resulting oils. Trials were carried out over a 3-year period on trees subjected to irrigation and grown under rainfed conditions. Vegetative tree response, as shoot elongation and trunk diameter, was evaluated. Yield per plant, fruit characteristics, oil quality indices (free fatty acid content, peroxide value, UV adsorption at 232 and 270 nm, total phenols, α-tocopherol content) were determined for both irrigated and non-irrigated treatments in the ‘on’ years 1997 and 1999 (6th and 8th year after planting, respectively).     

Utility of RAPD,ISSR, IRAP and REMAP markers for the genetic analysis of Citrus spp.

The application and informativeness of RAPD, ISSR, IRAP and REMAP markers to study the genetic diversity and relationship among the Citrus and its relative genotypes were investigated. High levels of polymorphism were observed for all four marker systems. The RAPD technique generated the highest number of polymorphic bands and average number of polymorphic band per assay unit. Average limit of the discriminating power was very close to its actual discriminating power of each marker. The highest and the lowest values of effective number of patterns were obtained from the marker REMAP (5.94) and RAPD (4.48), respectively. Correlation between the genetic similarities matrices were estimated from all four markers using the Mantel matrix correspondence test, and results showed significant correlations among the RAPD, ISSR, IRAP and REMAP similarity matrices. The highest correlations were found comparing RAPD and ISSR markers, whereas RAPD and REMAP (r = 0.143) markers were poorly correlated. To assess the usefulness of the overall information provided by these marker data for establishing phylogenetic relationships and Citrus germplasm classification, cluster analysis was performed. All four techniques, solely or in combination, discriminated the genotypes very efficiently and generated a high similarity in dendrogram topologies, although some differences were observed. The linkage analysis was conducted based on the segregation of 38 RAPD, 13 ISSR, 19 IRAP and 9 REMAP loci in 96 progeny of an intergeneric cross Citrus sinensis and Poncirus trifoliata. Among the 81 studied loci 65 loci distributed on five linkage groups. Comparing the result obtained with RAPD, ISSR, IRAP and REMAP markers in this study, IRAP and REMAP proved to be as a reliable molecular marker for fingerprinting, mapping and diversity study of Citrus and its relatives.     

Agronomic evaluation of seedlings from crosses between the main Spanish olive cultivar ‘Picual’ and two wild olive trees

Summary

Until recently, olive breeding programmes have been based exclusively on cultivated olive germplasm (Olea europaea subsp. europaea var. europaea), in contrast to breeding in other fruit crops where the use of wild relatives has been widely reported. In this study, ten agronomic traits were evaluated in two progenies derived from controlled crosses between the Spanish olive cultivar ‘Picual’ (female) and two wild (O. europaea subsp. europaea var. sylvestris) genotypes (males). The results of this evaluation were compared with the progenies of crosses between ‘Picual’ (female) and the three cultivars, ‘Sikitita’, ‘Meski’, and ‘Zaity’ (male). The two ‘Picual’ × wild genotype progenies had the highest mean values for vigour traits (i.e., tree height and trunk diameter). Progenies from both ‘Picual’ × wild genotype crossess also showed the highest percentages of trees (53.8% and 37.5%) with a short juvenile period, compared to the progenies from crosses between ‘Picual’ and each of the three cultivars ‘Sikitita’, ‘Meski’, and ‘Zaity’ (0%, 5%, and 3.6%, respectively). Significant differences were also observed between progenies in fruit traits as well as in oil contents and fruit weights. Progenies from both crosses with wild olive showed lower values for the fruit traits evaluated than the cultivated olive progenies. However, significant improvements were achieved compared to fruit traits in the wild parents. The implications of these results for the future use of wild germplasm in olive breeding programmes are discussed.     

Molecular characterization and genetic diversity of Prunus rootstocks

Twenty microsatellite primer pairs, previously developed in peach, were used to characterize and to explore genetic relationships among 44 clones, representing three groups of rootstocks defined as: (1) Peach-based rootstocks (Prunus dulcis × P. persica, P. persica × P. davidiana); (2) Myrobalan-Marianna plums (P. cerasifera and interspecific hybrids having P. cerasifera as a parent); and (3) Slow growing plums (P. insititia, P. domestica, and P. domestica × P. spinosa). Eighteen SSR markers, from the 20 initially used, were able to amplify polymorphic products for the Peach-based rootstocks and 13 common markers gave also polymorphism for the Myrobalan-Marianna and Slow growing plums groups. The Dice coefficient of similarity was calculated between all pairs of accessions and their genetic similarity represented by a principal coordinate analysis. The genetic diversity detected among the 44 clones studied divided them in three groups, which are in agreement with their current taxonomic classification and their morphological characteristics. A set of three microsatellites (BPPCT001, CPPCT022 and UDP98-407) can distinguish between all the clones analyzed. The analysis within groups reveal another two sets of three SSR to distinguish between the clones from the Peach based rootstocks and the Myrobalan-Marianna plums, respectively, and only a single SSR is needed to distinguish within the clones from the Slow growing plums group. These results demonstrate the high potential of the SSR analysis for peach rootstock identification and studies of diversity in Prunus species.     

Genetic diversity and relationships of wild and cultivated olives at regional level in Spain

Genetic diversity and relationships between local cultivars and wild olive trees from three important Spanish olive-growing regions, Andalusia (South), Catalonia and Valencia (from Eastern Mediterranean Coastal area), were studied by means of eight SSR loci. Distinct allelic composition and heterozygosity levels were found in wild olive populations and cultivars. The observed patterns of genetic variation revealed: a) the independent clustering of Andalusian wild olives in a separate gene pool, b) the belonging of wild populations and most cultivars from Catalonia to another gene pool, c) the joined clustering of Andalusian and a set of Valencian cultivars in a third gene pool, and d) clustering of wild individuals from Valencia to the three different gene pools. These results suggest that wild populations of Andalusia may represent true oleasters, the ones from Catalonia may be feral forms derived from cultivar seed spreading, while the population of Valencia seems to be the most admixed one. The significant differentiation between Andalusian and most Catalonian cultivars is indicating an independent selection of olive cultivars in the two regions. The detection of a certain wild genetic background in some Catalonian and Valencian cultivars and the similarity found between wild and cultivated forms may suggest the use of local wild trees in olive domestication. The proposed scenario for the development of olive cultivars in Andalusia includes an empirical selection of outstanding local wild genotypes followed by various generations of crosses and various replanting campaigns, as well as possible introductions of ancestral cultivars. Therefore, our findings would lead us to support the hypothesis that the current diversity found in Spanish olive cultivars may be regionally differentiated and due to both, autochthonous and allochthonous origin. The information obtained in this work gives insights into the genetic resources of the main olive producing country, demonstrating that wild olive populations and local cultivars both represent potential sources of useful variability for olive breeding programs.     

RAPD markers reveal polymorphism among some Iranian pomegranate (Punica granatum L.) genotypes

In this study RAPD markers were used to determine the diversity level among 24 Iranian pomegranate genotypes. One hundred decamer random primers were used for PCR reactions, among which 16 showed reliable polymorphic patterns. These primers produced 178 bands, of which 102 were polymorphic. Cluster analysis of the genotypes was performed based on data from polymorphic RAPD bands, using Jaccard's similarity coefficient and UPGMA clustering method. The highest and lowest similarities detected between genotypes were 0.89 and 0.29, respectively. At a similarity of 60%, the genotypes were divided into four sub-clusters. Cophenetic correlation coefficient between similarity matrix and cophenetic matrix of dendrogram was relatively high (r = 0.9) showing the goodness of fit of the dendrogram. RAPD markers showed to be a useful tool for studying the genetic diversity of pomegranate.     

Sap flow measurements in young olive trees (Olea europaea L.) cv. Chétoui under Tunisian conditions

Measurements of sap flux were carried out from May 2003 to March 2004 on 6 year-old irrigated olive trees of cultivar Chétoui cultivated at 6 m × 6 m spacing in Mornag (36.5°N, 10.2°E), Northern Tunisia. The aim of the research is to evaluate the sap flux technique for its applicability with young olive trees and to estimate their water consumption under field conditions. Three thermal sensors were implanted in the trunk of three olive trees following to North (N), South-East (SE) and South-West (SW) directions. Data were analyzed following to the procedure of Do and Rocheteau (2002b) that derives from Granier (1985). In this paper, data on probe calibration, wood conductive section estimation and sap flux spatial-variability are presented and discussed. Relationships between sap flux measurements, climate and soil water status have been investigated. Results show that sap flux values vary with sensor position, soil water content and climate demand. Good agreements between sap fluxes and global radiation and reference evapotranspiration measurements were observed. Some variations were recorded under water shortage conditions. Maximum and minimum daily fluxes of 4.5 l and 41.0 l per tree were found in February 2003 and in August 2003, respectively. Maximum transpiration represented only 53% of the crop evapotranspiration as determined by the F.A.O. method.     

Genetic diversity of Dianthus accessions as assessed using two molecular marker systems (SRAPs and ISSRs) and morphological traits

Dianthus chinensis, Dianthus barbatus and Dianthus superbus are members of the Caryophyllaceae and are grown widely as ornamental plants. Information about relative genetic relationships can facilitate breeding programs. Here, we have compared two polymerase chain reaction (PCR)-based systems (sequence-related amplified polymorphisms (SRAPs) and inter-simple sequence repeats (ISSRs)) and morpological trait measurements for their relative effectiveness in estimating the genetic diversity found between 22 Chinese pink (D. chinensis) inbred-lines, one accession of D. barbatus and one accession of D. superbus. Interspecific differences were readily detected but the markers were less reliable in distinguishing the accessions according to their region of origin or in separating the wild species from the cultivars. Morphological traits were found to be the least effective genetic markers. The relative effectiveness of the three systems as markers for genetic diversity was concluded to be SRAP > ISSR > morphological traits, but the combined data from ISSR + SRAP analyses was superior to all three. The information generated by the SRAP marker system correlated more closely with morphological variability than did the ISSR marker system. The morphological markers of plant height/crown size ratio, lower leaf length, ovary shape index and calyx length showed strong correlations with the genetic diversity index (GD_ij, PPB(II) and PSB) as generated by the percentage of polymorphic bands and percentage of special bands of the PCR-based markers.     

Microsatellite marker-based identification of mother plants for the reliable propagation of olive (Olea europaea L.) cultivars in Australia

Summary

Olive production in Australia has continued to increase in recent years, however there remains a high degree of confusion on the genetic identities of the cultivars being grown. In the present study, seven microsatellite (simple sequence repeat; SSR) loci were used to identify a set of 53 olive tree samples from different sources. The microsatellite DNA profiles of all 53 tree samples, including seven unknown trees, were compared with the SSR profiles of 14 reference olive cultivars. A total of 60 fragments (alleles), averaging 8.57 alleles per microsatellite locus, were amplified. High average values were found for the observed heterozygosity, the expected heterozygosity, and the polymorphic information content (0.73, 0.74, and 0.72, respectively). While all seven microsatellite markers proved useful for characterisation and identification purposes, a combination of three SSR primer pairs (DCA9, DCA18, and EM030) was sufficient to distinguish all 53 olive samples. The microsatellite allelic profiles allowed the 53 tree samples to be grouped into 23 genotypes. The allelic profiles of 14 of these genotypes matched with their reference cultivars, while the genetic identities of the remaining nine genotypes could not be confirmed. Some of these unknown genotypes may have been derived from feral olive trees, or were due to mislabelling and/or planting errors among Australian olive cultivars. Our results confirm the usefulness of microsatellite markers as a tool for cultivar differentiation and identification, and indicate the need for reliable identification of mother plants for commercial propagation.     

Compositional quality of virgin olive oils from cultivars introduced in Tunisian arid zones in comparison to Chemlali cultivars

This work is an evaluation of the quality of three introduced European olive cultivars Arbequina, Coratina, and Koroneiki grown under a biological agricultural system in the arid zone of Sfax in Tunisia compared to Chemlali cultivars. Various olive parameters were analyzed, such as ripening index, pulp/stone (P/S) ratio and oil content. We have considered the regulated physicochemical analytical parameters, fatty acids and minor compounds for olive oil. The most of the quality indices and fatty acid composition showed significant variations among olive cultivars. The Koroneiki, Coratina and Chemlali Zarzis cultivars had highest values of oleic acid (62.7%) (76.8%, 75.8% and 73.9%, respectively), Arbequina, had lowest value of oleic acid and similar to Chemlali Sfax. Coratina was noteworthy for its higher content of phenolic compounds (287.8 mg kg⁻¹), oil content (42.4%) and P/S ratio (4.7%). Although their low phenol contents, autochthonous cultivars presented high contents of α-tocopherol (577.8 and 434.6 mg kg⁻¹ for Chemlali Boughrara and Chemlali Zarzis, respectively) except for Chemlali Sfax. In conclusion, European cultivars seem to have adapted to the area studied according to their oil biochemical composition while the increase observed in their α-tocopherol levels. The cultivar Arbequina showed a similar composition of Chemlali Sfax.