Genetic Structure in Tunisian Apricot, Prunus armeniaca L., Populations Propagated by Grafting: A Signature of Bottleneck Effects and Ancient Propagation by Seedlings

In order to give insights into the origin and historical selection process of Tunisian apricot propagated by grafting, 31 cultivars from three areas presenting contrasting ecological conditions – Kairouan, Testour and Ras Jbel were compared to cultivars from Europe, North America, North Africa, Turkey, Iran and China, using 234 AFLP markers. The phenetic analysis allowed to distinguish 5 clusters, the four previously defined groups: – ‘diversification’, ‘geographically adaptable’, ‘continental European’ and ‘Mediterranean’ – groups and the Tunisian one. The partitioning of genetic diversity within and between cultivar groups assessed according to the Bayesian approach and assuming Hardy–Weinberg equilibrium, showed a loss of 21.81–38.49% of genetic diversity in Tunisian apricot compared to Mediterranean and diversification groups, respectively. Genetic variation occurred within Tunisian subgroups rather than among (F_ST = 0.060) evidencing a narrow genetic pool. Mediterranean and Tunisian groups were the least differentiated. Comparing them, 24 AFLP fragments discriminated the Mediterranean group from the Tunisian group but most of them where also shared by the other groups. Strongly differentiated gene pool and low genetic diversity are probably the result of bottleneck effects linked to the occurrence of propagation by seedlings rather than by grafting during the introduction periods in the North and the Centre of Tunisia. This study points at the propagation by seedlings as an important factor which should be taken into account to understand the evolution of apricot in South Mediterranean areas.     

Inter-Simple Sequence Repeat fingerprints to assess genetic diversity in Tunisian fig (Ficus carica L.) germplasm

The genetic diversity of 18 Tunisian fig cultivars was investigated at the DNA level using the Inter Simple Sequence Repeat (ISSR) associated with the Polymerase Chain Reaction (PCR). Using a set of primers, the most informative ones were selected that were characterized by an important Resolving power value of 29.6. A total of 47 discernible fragments were scored from samples, with a mean of 11.7 fragments per primer. The 90.4% of sample that were polymorphic were scored as molecular markers to examine the Tunisian fig germplasm polymorphism at DNA level. A large genetic diversity as related to ISSR patterns was found within the local Tunisian fig germplasm. An UPGMA dendrogram exhibits the unstructured variability in this crop. Moreover, the principal component analysis shows that the observed diversity was typically continuous. Our data provide a large number of ISSR markers that are useful in the fingerprinting of Ficus carica L. cultivars, and in the understanding of the genetic relationships among these accessions.     

Comparative Analysis of Genetic Diversity in Two Tunisian Collections of Fig Cultivars Based on Random Amplified Polymorphic DNA and Inter Simple Sequence Repeats Fingerprints

This study characterized the genetic diversity of 18 Tunisian fig cultivars using random amplified polymorphic DNA (RAPD) and inter simple sequence repeats (ISSR). Both random and ISSR primers tested generated a total of 116 RAPD and 47 ISSR markers. Considerable genetic variation was observed among fig cultivars sampled from two regional Tunisian collections with an average diversity of 4.57. RAPD and ISSR banding patterns and genetic distances values reflected the high level of diversity among the collections and lower variability between the two collections. The correlation between the RAPD and ISSR similarity matrices computed for the 153 pairwise comparisons among the 18 varieties was lower and significant. An analysis of molecular variance showed that 92% of the total genetic diversity resided within collections, whereas only 8% between collections. The results indicated that in the local fig germ plasm the information provided by RAPD and ISSR is not analogous, most likely as a consequence of the fact that the two classes of markers explore, at least in part, different portions of the genome.     

Genetic diversity in Tunisian perennial ryegrass revealed by ISSR markers

The genetic diversity in Tunisian perennial ryegrass (Lolium perenne) was examined by the help of inter-simple sequence repeats (ISSR). Starting from eighteen accessions, a large number of polymorphic ISSR markers were currently generated using appropriate primers (a total of 136, which average of 12.6 polymorphic bands/primer). These markers were considered to estimate the genetic distance among accessions and to draw phylogenetic trees. Our data provide evidence of a high degree of genetic diversity in Tunisian ryegrass. In addition, both cultivars and wild types present a high degree of divergence suggesting a complex domestication process in this crop. Moreover, spontaneous populations of Tunisian ryegrass have been identified as important ecotypes that are suitable in selection programs to improve grasslands.     

Genetic diversity of cowpea [<Emphasis Type="Italic">Vigna unguiculata</Emphasis> (L.) Walp.] in four West African and USA breeding programs as determined by AFLP analysis

Cowpea is an important grain legume and hay crop of many tropical and subtropical regions, especially in the dry savanna region of West Africa. The cowpea gene pool may be narrow because of a genetic bottleneck during domestication. Genetic variation within specific breeding programs may be further restricted due to breeding methods, ‘founder effects’ and limited exchange of germplasm between breeding programs. Genetic relationships among 60 advanced breeding lines from six breeding programs in West Africa and USA, and 27 landrace accessions from Africa, Asia, and South America were examined using amplified fragment length polymorphism (AFLP) markers with six near infrared fluorescence labeled EcoRI + 3/1bases/MseI + 3/1bases primer sets. A total of 382 bands were scored among the accessions with 207 polymorphic bands (54.2%). Despite a diverse origin, the 87 cowpea accessions shared a minimum 86% genetic similarity. Principal coordinates analysis showed clustering of breeding lines by program origin, indicating lack of genetic diversity compared to potential diversity. Accessions from Asia and the Americas overlapped and were distinct from West African breeding lines, indicating that germplasm from Asia and the Americas have common origins outside West Africa. US and Asian breeding programs could increase genetic variability in their programs substantially by incorporating germplasm from West Africa, while national programs in West Africa should consider introgression of Asian germplasm and germplasm from other parts of Africa into their programs to ensure long-term gains from selection.     

SSR Diversity of Vegetable Soybean [<Emphasis Type="Italic">Glycine max</Emphasis> (L.) Merr.]

Edamame [Glycine max (L.) Merr.] is a type of soybean selected for fresh or frozen vegetable use at an immature stage. Since edamame has a similar protein content, milder flavor, nuttier texture, and is easier to cook when compared to grain soybean, it is being promoted as a new vegetable for global consumption. Global production will require breeding programs for local adaptation; however, limited research has been published on genetic diversity of edamame varieties for the assessment of genetic resources. Simple sequence repeats (SSRs) were used to study the genetic diversity among 130 accessions, including edamame cultivars and landraces from Japan, China and the US, and also the new breeding lines in the US. Although it is assumed that elite edamame cultivars would have narrow genetic diversity, seventeen SSRs detected polymorphism to distinguish 99 of the 130 accessions. The cluster analysis generated nine clusters and 18 outliers. Genetic diversity within Japanese edamame was lower than that within Chinese vegetable soybean accessions (maodou), even though only 10 Chinese maodou were analyzed compared to 107 Japanese edamame. Cluster analysis revealed that the patterns of SSR diversity in edamame can generally distinguish maturity classes and testa color. We concluded that Japanese edamame have a narrow genetic base different from others and that SSRs can describe the patterns of genetic diversity among the elite vegetable soybean.     

Intra-specific DNA polymorphism in pineapple (<Emphasis Type="Italic">Ananas comosus</Emphasis> (L.) Merr.) assessed by AFLP markers

Pineapple (Ananas comosus (L.) Merr.) cultivars, often derived from somatic mutations, are propagated vegetatively. It has been suggested by isozyme data that there is little genetic variation among Smooth Cayenne cultivars. A thorough investigation of the genetic variation within the cultivated speciesAnanas comosus, particularly among commercial cultivars, will provide critical information needed for crop improvement and cultivar protection. One-hundred and forty-eight accessions ofA. comosus and 14 accessions of related species were evaluated with AFLP markers. The average genetic similarity ofA. comosus was 0.735 ranging from 0.549 to 0.972, suggesting a high degree of genetic variation within this species. With AFLP markers, discrete DNA fingerprints were detected for each commercial cultivar, breeding line, and intra-specific hybrid. Self-incompatibility, high levels of somatic mutation, and intraspecific hybridization may account for this high degree of variation. However, major cultivar groups of pineapple, such as Cayenne, Spanish, and Queen, could not be distinctively separated. These cultivar groups are based on morphological similarity, and the similar appearance can be caused by a few mutations that occurred on different genetic background. Our results suggest that there is abundant genetic variation within existing pineapple germplasm for selection, and discrete DNA fingerprinting patterns for commercial cultivars can be detected for cultivar protection. The genetic diversity and relationships of fourAnanas species are also discussed.     

RAPD and ISSR fingerprinting in cultivated chickpea (<Emphasis Type="Italic">Cicer arietinum</Emphasis> L.) and its wild progenitor <Emphasis Type="Italic">Cicer reticulatum</Emphasis> Ladizinsky

Detection of genetic relationships between 19 chickpea cultivars and five accessions of its wild progenitor Cicer reticulatum Ladizinsky were investigated by using RAPD and ISSR markers. On an average, six bands per primer were observed in RAPD analysis and 11 bands per primer in ISSR analysis. In RAPD, the wild accessions shared 77.8% polymorphic bands with chickpea cultivars, whereas they shared 79.6% polymorphic bands in ISSR analysis. In RAPD analysis 51.7% and 50.5% polymorphic bands were observed among wild accessions and chickpea cultivars, respectively. Similarly, 65.63% and 56.25% polymorphic bands were found in ISSR analysis. The dendrogram developed by pooling the data of RAPD and ISSR analysis revealed that the wild accessions and the ICCV lines showed similar pattern with the dendrogram of RAPD analysis. The ISSR analysis clearly indicated that even with six polymorphic primers, reliable estimation of genetic diversity could be obtained, while nearly 30 primers are required for RAPD. Moreover, RAPD can cause genotyping errors due to competition in the amplification of all RAPD fragments. The markers generated by ISSR and RAPD assays can provide practical information for the management of genetic resources. For the selection of good parental material in breeding programs the genetic data produced through ISSR can be used to correlate with the relationship measures based on pedigree data and morphological traits to minimize the individual inaccuracies in chickpea.     

Genetic inheritance analysis of four enzymes in date-palm (Phoenix dactylifera L.)

The genetic inheritance of four date-palm enzymes hasbeen carried out in 29 Tunisian cultivars. Our data provide evidenceof five polymorphic genes involving 12 alleles. It has been assumedthat either Pgm or Shdh are specified by three different alleles,while Got-1, Got-2 and Pgi-2 loci are biallelicgenes. On the other hand, analysis throughout progenies of twocontrolled crosses support significantly these hypothesis. In thewhole, results are in agreement with reports that describeddate-palm isozymes. Nevertheless, we assume that other alleleare involved in Pgm gene. Also, isozymes controlled by Got-2locus are of a dimeric structure. Opportunely, it can be establishedthat the resultant isoenzymes would constitute polymorphic markerssuitable to analyze the genetic diversity in date-palm.     

Genetic diversity changes and relationships in spring barley (<Emphasis Type="Italic">Hordeum vulgare</Emphasis> L.) germplasm of Nordic and Baltic areas as shown by SSR markers

To examine changes in the level of and pattern in variability in 197 Nordic and Baltic spring barley cultivars over time we used 21 mapped barley simple sequence repeats (SSRs). A total number of 191 alleles were found from 22 SSR loci. The number of alleles per locus ranged from 2 to 23, with average of 8.63 107 alleles were rare (frequency <0.05) among the cultivars and only one allele was frequently observed (frequency >0.95). The gene diversity between loci in Nordic and Baltic material varied between 0.033 and 0.891. Average gene diversity was 0.623. The SSR data separated two-rowed and six-rowed cultivars. According to analysis of molecular variance (AMOVA) differentiation in two-rowed vs six-rowed accounted for 23.6% of the total variation. Overall no significant decrease of average gene diversity over time could be found. However, differences were observed when spring barleys from northern (north of ∼58°) and southern (south of ∼58°) parts of the Nordic and Baltic area were compared. For the southern ecogeographical region significant decrease of genetic diversity was observed in the middle of the 20th century, whereas no significant changes in the northern part were found. We found larger differentiation between modern and old cultivars in the South compared to the ones in North parts of the region. The magnitude of changes in genetic diversity differed also with the country of origin. Danish cultivars had a significant decrease in diversity in the middle of century, whereas changes in Finland, Norway and Sweden were not significant.     

The development of a new approach for establishing a core collection using multivariate analyses with tulip as case

A methodology is developed for the establishment of a core collection based on phenotypic data. A case is worked out for the tulip cultivar collection of the Hortus Bulborum, consisting of approximately 1000 cultivars. The methodology is based on cluster analysis of phenetic characters, selection of cultivars from the resulting clusters according to a combination of the proportional and genetic diversity dependent strategies, optimisation of the selection, and validation by means of principal component analysis and by the diversity indices of Nei and Shannon & Weaver. A core collection with 104 cultivars (approx. 10%: CORE-1) did not give a sufficient representation of the existing diversity. A set of 152 cultivars (approx. 15%: CORE-2) showed a far better representation. The variation in resistance levels for Fusarium is almost completely represented in CORE-2. Although a set of 200 cultivars (approx. 20%: CORE-3) reached a nearly complete representation of the total diversity, a better representation of the resistance levels was not achieved. The newly presented methodology for defining a core collection appears to be a useful approach. The included steps for optimising and validating the choice of accessions makes it a reliable and broadly applicable method.     

RAPD and SSR markers for characterization and identification of ancient cultivars of <Emphasis Type="Italic">Olea europaea</Emphasis> L. in the Emilia region,Northern Italy

The Emilia region (Northern Italy) is characterised by the occurrence of microclimates that permit olive growing. The presence of the species, albeit sporadic, in these territories for several centuries as a fruit crop is well documented, by both archaeological and written testimony, and by a large number of plants well over a century old, located in particular sites, favourable for growth and development of the tree. Olive genetic diversity was studied using RAPD and SSR techniques, on plants growing in the Emilia territory (Reggio Emilia and Parma provinces). For genotype identification comparisons were made with 8 cultivars, some of which from Central Italy. Screening was obtained analysing patterns produced by 20 RAPD primers and 3 SSR primers, developed by other authors; the primers and we were able to discriminate olive cultivars with a sufficient degree of reliability. The dendrograms obtained from the analysis show the genetic relationship among accessions present in the Parma-Reggio Emilia district. Our results demonstrated the reliability of RAPDs and SSRs to identify all studied olive cultivars and to reveal the degree of their relatedness to each other. The analysis also reveals the presence of an interesting amount of genetic diversity among the studied individuals.     

Studying the extent of genetic diversity among Gossypium arboreum L. genotypes/cultivars using DNA fingerprinting

Genetic diversity is an area of concern for sustaining crop yield. Information on genetic relatedness/diversity among Gossypium arboreum L. cultivars/genotypes is scanty. We have used random amplified polymorphic DNA (RAPD) analysis to assess the genetic divergence/relationship among 30 genotypes/cultivars of G. arboreum. Of 45 primers surveyed, 63% were polymorphic. Out of the total number of loci amplified, 36% were polymorphic. The calculated genetic similarity between the cultivars/genotypes was in the range of 47.05–98.73%. Two genotypes, HK-244 and Entry-17, were the most distantly related. The average genetic relatedness among all the genotypes was 80.46%. However, most of the cultivated varieties showed a close genetic relationship, indicating a narrow genetic base in comparison to the non-cultivated germplasm. The calculated coefficients were used to construct a dendrogram using the unweighted pair group of arithmetic means (UPGMA) algorithm, which grouped the genotypes/cultivars into two major and three smaller clusters. The study is the first comprehensive analysis of the genetic diversity of G. arboreum germplasm and identifies cultivars that will be useful in extending the genetic diversity of cultivated varieties and future genome mapping projects.     

A study of genetic diversity among Indian bread wheat (Triticum aestivum L.) cultivars released during last 100?years

Genetic diversity was examined in a collection of 263 Indian bread wheat (Triticum aestivum L.) cultivars using 90 SSR markers. These cultivars were classified into Group I (pre-green revolution cultivars) and Group II cultivars (post-green revolution cultivars). SSR markers were also classified into Set-I SSRs (42 random genomic SSRs) and Set-II SSRs (48 SSRs associated with QTLs for grain weight). The SSRs belonging to Set-II exhibited relatively low level of polymorphism, suggesting that the SSRs associated with QTL for grain weight in wheat were probably under selection pressure during wheat breeding. AMOVA indicated that proportion of the variation within each of the two groups of cultivars accounted for most (87.59%) of the molecular variance, while the variation between the two groups of cultivars accounted for only 12.41% of the variance. The estimates of the average number of alleles/locus and gene diversity due to each of the two sets of SSRs suggested increase in overall genetic diversity after green revolution in Indian bread wheat cultivars. Differences were also observed in genetic diversity among cultivars from each of the six wheat growing agro-climatic zones of India. However, decade-wise analysis of genetic diversity among the post-green revolution cultivars indicated a progressive decline in genetic diversity, thus suggesting a need for involving diverse exotic, synthetic and winter wheat germplasm in Indian wheat breeding programs.     

Application of random amplified polymorphic DNA to study genetic diversity in Paspalum scrobiculatum L. (Kodo millet, Poaceae)

Summary Genetic diversity and patterns of geographic variation among collections of Paspalum scrobiculatum (kodo millet) and P. polystachyum were studied using molecular markers generated through the random amplified polymorphic DNA (RAPD) method. A high level of polymorphism in RAPD markers was observed among the individual accessions, demonstrating the high genetic diversity of the crop. The markers obtained from the RAPD method were analyzed with the cluster analysis, principal coordinates and minimum spanning tree methods. Three major groups were resolved, one representing the African accessions, and two for the Indian accessions. The accessions of the north African kodo millet and P. polystachyum (considered conspecific with P. scrobiculatum) were quite distinct. The Australian kodo millet showed higher affinity to the African types. The study demonstrated that the RAPD technique can be applied to resolving degrees and patterns of genetic variation at the population and species levels, identifying cultivars, and defining gene pools of this crop.     

Genetic diversity of landraces of wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) from hilly areas of Uttaranchal,India

Seed samples of 27 landraces of wheat were collected from farmers’ fields of hilly areas of Himalaya in Uttaranchal state of India during April 2004. Genetic diversity among 41 genotypes (cultivars and landraces of wheat) was studied using morphological traits, microsatellite markers and SDS-PAGE of HMW-GS. The dendrogram and PCA (Principal Component Analysis) based on morphological data clearly separated landraces of wheat from cultivars. In the dendrogram based on microsatellite markers data all the wheat cultivars released after the introduction of high yielding dwarf wheat varieties from CIMMYT, used in this study, were grouped separately with the exception of NP4. The pre-green revolution indigenous varieties grouped with landraces suggesting that the same had been probably developed through selection among landraces in India. The landraces had higher diversity for HMW-glutenin subunits coded by Glu-B1, with distinct subunit combinations 6 + 8, 7 + 9, 13 + 16, than within the wheat cultivars analyzed. Most of the landraces except IITR10 and IITR14 are clearly distinct from the indigenous and modern wheat cultivars released in India in the 20th century. More than half of the landraces were heterogeneous mixture of plants with different glume color, awnness, grain color and HMW-GS profile and hence need purification through single plant selection. Some of the landraces with resistance to yellow rust and powdery mildew and distinct HMW-GS subunits can be used in appropriate breeding programs. It will be desirable to conserve and protect the landraces as geographical indications of Uttaranchal.     

Collection of Lupinus angustifolius L. Germplasm and Characterisation of Morphological and Molecular Diversity

Lupinus angustifolius L. is a Mediterranean species, domesticated in the 20th century, representing an important grain legume crop in Australia and other countries. This work is focused on the collection of wild germplasm and on the characterisation of morphological and molecular diversity of germplasm accessions. It reports the collection of 81 wild L. angustifolius accessions from the South and Centre of Portugal, available at the ‘Instituto Superior de Agronomia Gene Bank’, with subsequent morphological and molecular characterisation of a selection of these and other accessions. A multivariate analysis of morphological traits on 88 L. angustifolius accessions (including 59 wild Portuguese accessions, 15 cultivars and 14 breeding lines) showed a cline of variation on wild germplasm, with plants from Southern Portugal characterised by earlier flowering, higher vegetative development and larger seeds. AFLP and ISSR molecular markers grouped modern cultivars as sub-clusters within the wider diversity of wild germplasm, revealing the narrow pool of genetic diversity on which domesticated accessions are based. The importance of preserving, characterising and using wild genetic resources for L. angustifolius crop improvement is outlined by the results obtained.     

Abundant genetic diversity in cultivated <Emphasis Type="Italic">Codonopsis pilosula</Emphasis> populations revealed by RAPD polymorphisms

Genetic diversity of seven cultivated populations of Codonopsis pilosula Nannf. from Longxi County, Gansu Province of China was estimated using randomly amplified polymorphic DNA (RAPD) markers. The 17 selected RAPD primers amplified 205 polymorphic bands out of a total of 235 (87.2%). Nei’s gene-diversity statistics and population differentiation parameters based on AMOVA analysis indicated that the cultivated C. pilosula populations remained a high level of genetic diversity with Hs = 0.299 and I = 0.450. A greater proportion of genetic diversity was found within (77%) rather than among (23%) the populations. In addition, we also detected that populations from different altitudes had a considerable genetic differentiation after 40 years of cultivation at the same site. Populations from higher altitude had lower genetic diversity than those from lower altitude. Our results suggested that irregular and sparse cultivation practices, i.e., random collecting, preserving, and planting seeds of the medicinal species without deliberate selection, might be an efficient way to conserve genetic resources of medicinal plants, in addition to their effective uses.     

Analysis of Genetic Diversity in Barley Cultivars Reveals Incongruence Between S-SAP,SNP and Pedigree Data

Accurate assessment of genetic similarity is important for plant breeding, germplasm enhancement and conservation of plant genetic resources. A comparative analysis of genome diversity among a group of six-rowed spring barley (Hordeum vulgare L.) cultivars was carried out using sequence-specific amplified polymorphism (S-SAP) and single nucleotide polymorphism (SNP), with the results compared to the kinship coefficients derived from the pedigree data. Mean pair-wise GS values were estimated to be 0.0957 ± 0.144 (Kinship), 0.491 ± 0.189 (SNPs), and 0.602 ± 0.098 (S-SAPs). S-SAP and SNP-based genetic similarity (GS) values were normally distributed but kinship values had a non-normal and skewed distribution. Pair-wise correlation of GS values were lowest for the S-SAP and the SNP matrices (r =; 0.040, p<0.230) and highest for the SNP and pedigree matrices (r =; 0.240, p < 0.001). Analysis of molecular variance (AMOVA) attributed about 90.4% of observed variation to the cultivars within each of the malting and feed groups. Variance component between malting and feed groups was 6.6% for both SNP and S-SAP data suggesting lack of a significant genetic differentiation along this agronomic division. The remaining 3% of variation was attributed to genetic diversity within cultivars. Although both DNA-based marker systems were able to differentiate all barley cultivars, significant difference were observed in the pattern of genetic relationships obtained by the two marker systems and the pedigree data.