Assessing Molecular Signature for Some Potential Date (Phoenix dactylifera L.) Cultivars from Saudi Arabia, Based on Chloroplast DNA Sequences rpoB and psbA-trnH

Phoenix dactylifera L. (date palm), being economically very important, is widely cultivated in the Middle East and North Africa, having about 400 different cultivars. Assessment of date cultivars under trading and farming is a widely accepted problem owing to lack of a unique molecular signature for specific date cultivars. In the present study, eight different cultivars of dates viz., Khodry, Khalas, Ruthana, Sukkari, Sefri, Segae, Ajwa and Hilali were sequenced for rpoB and psbA-trnH genes and analyzed using bioinformatics tools to establish a cultivar-specific molecular signature. The combined aligned data matrix was of 1147 characters, of which invariable and variable sites were found to be 958 and 173, respectively. The analysis clearly reveals three major groups of these cultivars: (i) Khodary, Sefri, Ajwa, Ruthana and Hilali (58% BS); (ii) Sukkari and Khalas (64% BS); and (iii) Segae. The economically most important cultivar Ajwa showed similarity with Khodary and Sefri (67% BS).The sequences of the date cultivars generated in the present study showed bootstrap values between 38% and 70% so these sequences could be carefully used as molecular signature for potential date cultivars under trading and selection of genuine cultivars at the seedling stage for farming

Molecular characterization and genetic relationships among most common identified morphotypes of critically endangered rare Moroccan species Argania spinosa (Sapotaceae) using RAPD and SSR markers

$\bullet$ The objective of this work is the molecular characterization of most common identified morphotypes of critically endangered rare Moroccan species Argania spinosa. $\bullet$ Eighteen RAPD markers and twenty SSR markers have been assayed in 38 argan tree accessions from the three most commonly identified morphotypes: oval, spherical and spindle fruit types. $\bullet$ A total of 140 polymorphic RAPD bands were detected out of 146 bands. The number of presumed alleles revealed by the SSR analysis ranged from 1 to 5 alleles per locus with a total number of 32 alleles identified. Results demonstrated an extensive genetic variability within the tested argan accessions. RAPDs presented a high level of polymorphism and greater information content than SSRs. $\bullet$ Our results could indicate that Vitellaria paradoxa is genetically closer to argan than Manilkara huberi. In addition, the correlation between the clustering based on RAPD and SSR markers were in general low. The observed clustering could be better explained according to geographic proximity than morphotype. For this reason the traditional morphological characterization of the argan accessions in morphotypes (according to fruit phenotype) seem not to be in correlation to the real genetic background (genotype) of this specie. The implications of these results in the creation of effective germplasm core collection in argan have been also discussed.Caractérisation moléculaire et relation génétique entre les morphotypes les plus connus d'une espèce marocaine rare et menacée, Argania spinosa (Sapotaceae) à l'aide de marqueurs RAPD et SSR. $\bullet$ L'objectif de ce travail consiste en la caractérisation moléculaire et l'étude des relations génétiques entre des morphotypes identifiés chez une espèce marocaine rare et menacée, l'arganier (Argania spinosa). $\bullet$ Pour cela, dix-neuf marqueurs RAPD et vingt marqueurs SSR on été testés dans 38 accessions d'arganier des trois morphotypes du fruit identifiés: ovale, sphérique et broche. $\bullet$ En utilisant la technique d'amplification au hasard de l'ADN (RAPD), 140 fragments polymorphes ont été détectés parmi les 146 amplifiés. Le nombre d'allèle révélé par l'analyse des marqueurs microsatelittes (SSRs) varie de un à cinq allèles par locus avec un total de 32 allèles identifiés. Les résultats démontrent une grande variabilité génétique au sein des accessions d'arganier testées. Les marqueurs RAPD présentent un taux de polymorphisme et une hétérozygotie attendue supérieurs aux SSR. $\bullet$ Les résultats indiquent également que Vitellaria paradoxa est plus proche génétiquement de l'arganier que Manilkara huberi. La corrélation entre les groupements observés dépend plus de la proximité géographique que de la classification selon la forme du fruit (morphotypes). La classification traditionnelle par morphotypes n'est donc pas en accord avec l'analyse génotypique chez cette espèce. L'apport de ces résultats pour la création d'une collection de référence chez cette espèce est discuté

Enhanced plantlet regeneration from cultured meristems in sprouting buds of saffron corms

4 pages.-- In: ISHS Acta Horticulturae 739: II International Symposium on Saffron Biology and Technology.When meristems dissected from dormant corms were used for micropropagation of saffron, low and heterogeneous morphogenic potential as well as high contamination levels have been found (Piqueras and Hernandez, 2004). With the aim to prevent the above mentioned inconvenient, shoot cultures have been initiated from meristems in sprouted saffron corms before flowering. These explants showed a significant lower contamination rate (less than 30%) when cultured in a culture medium composed of QL salts, MS vitamins and 30 g/L sucrose solidified with 7.5 g/L of agar. To study the morphogenic response induced by different cytokinin types and concentrations (BAP, 2iP and TDZ), the explants were cultivated in medium supplemented with them for six weeks. After this period, the number of new shoots per initial explant, their length and quality were recorded and used as parameters for the selection of the most effective cytokinin level for multiplication. By using this new explant for the initiation of axillary shoot cultures in saffron a new micropropagation procedure has been developed suitable for the clonal propagation or ex situ germplasm conservation of selected genotypes.Peer reviewe

Karyotype characterization of Argania spinosa (L.) Skeel (Sapotaceae)

3 pages, 1 figure.Argania spinosa (L.) Skeel is an endemic species from Southwest Morocco being the unique representative of the tropical Sapotaceae in this area. The cytology of this species is poorly known in spite of its great socio-economical and ecological interest in these arid and semi-arid zones. The objective of this work is to characterize the karyotype of A. spinosa species in somatic cells from root tips. Samples analyzed showed a karyotype constituted for ten pairs of chromosomes (2n = 2x = 20) and the putative karyotype proposed has been of four submetacentric and six metacentric pairs. The four submetacentric pairs were the longest with a mean total length between 1.14 and 1.69 μm and the total length of six metacentric pairs were between 0.59 and 1.03 μm.This work has been supported by grant A/6210/06 “Caractérisation biochimique et génomiques des morphotypes de l´arganier (Argania spinosa L. Skeels)” from the AECI (Agencia Española de Cooperación Internacional) of Spain.Peer reviewe

Molecular characterization using SSR markers and biochemical analysis of Moroccan and Spanish argan [Argania spinosa (L.) Skeels] ecotypes under water stress and rewatering

An integrated molecular and biochemical approach was carried out to compare Moroccan and Spanish argan [Argania spinosa (L.) Skeels] ecotypes. Genetic diversity of seedlings was evaluated using Simple Sequence Repeat (SSR) markers. In addition, water stress was imposed by holding water and enzymatic peroxidase (POX), superoxide dismutase (SOD), catalase (CAT) activities and total proteins in well-watered and water-deficient conditions were evaluated. The molecular characterization of the two ecotypes showed that the number of presumed alleles revealed by the SSR analysis ranged from 1 to 32 alleles per locus, with a mean value of 13 alleles and a mean heterozygosity value of 0.44. The assayed SSRs showed that most individuals were clustered according to their ecotype with a similar diversity level between Moroccan and Spanish ecotypes. Enzymatic activities in response to drought stress showed increase in POX and SOD activities for both ecotypes together with total protein content, but showed no significant differences in CAT activity. A potential tolerance to drought stress with increased levels of SOD activity in Spanish ecotypes grown in inland area of Alicante compared with Moroccan ecotypes grown in Essaouira, which is a coastal area, was also reported. In addition, the recovery period seemed to be sensed by argan plants as a new stress situation, as observed by the response of POX and CAT activities, suggesting an imbalance in the HO metabolism. Our results suggestd an effect of geographic proximity and population isolation and/or breeding system on molecular phenotypes and responses to drought stress of the studied argan ecotypes. However, further investigations are needed to verify the induced differences.This study has been supported by projects: “Selection and characterization of drought resistant almond cultivars from the Mediterranean basin with high nutraceutical values” (Nut4Drought) from ARIMNET-2 European Program and “Breeding stone fruit species assisted by molecular tools” from the Seneca Foundation of the Region of Murcia (19879/GERM/15

Valorization of saffron (Crocus sativus L.) stigma as a potential natural antioxidant for soybean (Glycine max L.) oil stabilization

Synthetic antioxidants are known for their efficiency to improve vegetable oil oxidative stability. But owing to their harmful effects on human health, edible oil industry is seeking for safe and healthy natural antioxidants. The present work was setup with the aim of improving soybean oil (SO) oxidative stability by using saffron (Crocus sativus L.) stigmas collected in Morocco. Saffron stigmas were used as a natural antioxidant at various concentrations (0.2, 0.3, and 0.6%) in soybean oil compared to tocobiol (0.3%) as a synthetic antioxidant (the positive control). Performances of such natural and synthetic antioxidants were evaluated by measuring oil basic quality indices under accelerated storage at 60 °C for 12 weeks. Such indices consisted of free fatty acids (FFA), peroxide value (PV), anisidine value (p-AV), total oxidation value (TOTOX), UV extinction coefficients (K232 and K270), fatty acids composition (FA), and iodine value (IV). The obtained data show that there were significant (p  0.9, were modeled through simple regressions. In conclusion, saffron stigmas with the different doses performed better than the positive control (tocobiol) regardless of the storage time. It could be concluded that saffron stigmas are a promising natural antioxidant, alternative to synthetic antioxidants, to enhance the oxidative stability of edible oils

Molecular characterization of almond cultivars and related wild species using nuclear and chloroplast DNA markers

6 pages.The genetic diversity of different accessions of several wild almond species including P. eleagnifolia, P. hausknechtii, P. scoparia and P. lycioides, endemic to Irano-Afghan plate, was investigated using nuclear and chloroplast DNA markers. In addition, five cultivated almond cultivars (Marta from Spain; Nonpareil and Mission from USA; Ferrangnes from France and Tuono from Italy) were included in the study. Diversity was analysed at the DNA level by means of 16 nuclear and 5 chloroplast pairs of primers flanking SSR sequences. In 45 accessions, the number of alleles per locus in nuclear microsatellites ranged from 7 to 16, and expected heterozygosity varied between 0.54 and 0.93 with average PIC value of 0.81. It showed that they represent rather polymorphic species. In the case of chloroplast SSR, the polymorphism observed was much lesser in agreement with higher level of conservation of the chloroplast DNA. Therefore, nuclear microsatellites have been the most abundant markers with a high polymorphism and level of transference across closely related species in comparison with the others. In addition, results indicated the high variability present in the S-locus. Hierarchical analysis using integrated nuclear and chloroplastic microsatellite data yielded best clustering with six logical clusters or groupings corresponding to the different subgenus and sections.Peer reviewe

Use of biotechnology for preserving rare fruit germplasm

15 pages.The application of recent biotechnological tools for conservation of rare fruit species from developing countries, including in vitro and hydroponic culture protocols, improved propagation techniques and molecular marker application, is described. Promising propagation methods include forcing germination of seeds, in-vitro protocols well adapted to these rare fruit species that allow the introduction, micropropagation and rooting of plant material, and developing hydroponic culture protocols that allow the early propagation of high-risk genotypes. In addition, the growth of seedlings in controlled environmental conditions in greenhouse and cold chamber provides a useful strategy for obtaining vigorously growing plants from seeds year round. A standard karyotyping protocol has been described working in several species as preliminary tool to start molecular (DNA) studies. In addition, different protocols for DNA isolation and quantification have been assayed in these rare fruit species. Molecular markers based on PCR amplification of the DNA have also become an essential tool for the characterization and conservation of these species. Regarding this PCR amplification of the DNA, two main strategies, RAPD (if the DNA sequence is unknown) and SSR markers (if the DNA sequence is known), have been assayed. These markers have been applied in the genetic characterization of this germplasm, the establishment of genetic relationships between cultivars and species, and the future construction of genetic maps of these rare fruit species. Additional advantages encouraging the utilization of these new technologies in breeding programs include the high levels of synteny between genomes of related species, and a well-established international network of cooperation among researchers.Peer reviewe

Application of recent biotechnologies in the conservation of rare fruit species from developing countries

International Symposium on Seed Enhancement and Seedling Production Technology -- 13 August 2006 through 19 August 2006 -- Seoul --Results of this work are from different international projects studying germ-plasm from rare fruit species in developing countries including the argan tree (Morocco), the date palm (Morocco) and several wild almond species (Iran). In this study, the application of recent biotechnological tools for conservation of these rare fruit species, including in vitro and hydroponic culture protocols, improved propagation techniques and molecular marker application, is described. Promising propagation methods include in vitro protocols well adapted to these rare fruit species that allow the introduction, micropropagation and rooting of plant material, and developing hydroponic culture protocols that allow the early propagation of high-risk genotypes. In addition, the growth of seedlings in controlled environmental conditions in greenhouse and cold chamber provides a useful strategy for obtaining vigorously growing plants from seeds year round. Molecular markers have also become an essential tool for the study and conservation of these species. Regarding the PCR amplification of the DNA, two main strategies have been applied, RAPD and SSR markers, in the genetic characterization of this germplasm, the establishment of genetic relationships between cultivars and species, and the future construction of genetic maps of these rare fruit species