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.     

37份龙眼种质资源亲缘关系的ISSR分析

本研究利用ISSR技术对37份龙眼种质资源进行遗传多样性检测。研究结果表明,从100条ISSR引物中筛选出7条重复性好,条带清晰的引物对37份龙眼品种基因组DNA进行扩增,共扩增出54条带,其中43条具有多态性,比率为79．6％。不同龙眼品种间遗传相似系数变幅为0．69～0．97,平均达0．83,说明ISSR标记能够揭示材料间较高的遗传多样性。UPGMA聚类结果表明,ISSR标记能将37份龙眼品种完全区分开,并能将来源于中国、越南和泰国的37份龙眼品种分别聚类到中国、越南和泰国三大品种群,说明龙眼品种资源的亲缘关系与地理因素有关,三个国家的龙眼品种之间存在较大的遗传差异。本研究结果将为为龙眼品种资源的研究利用提供参考。     

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.     

Characterization of Some Italian Common Bean (<Emphasis Type="Italic">Phaseolus Vulgaris</Emphasis> L.) Landraces by RAPD,Semi-random and ISSR Molecular Markers

Randomly amplified polymorphic DNA (RAPD), inter-simple sequence repeat (ISSR) and a semi-random PCR system were used to analyze the genetic diversity of 16 Italian common bean landraces and their relationship to four commercial cultivars. Of the primers tested, 8 ISSR, 6 RAPD and 7 semi-random primers produced polymorphic and reproducible DNA fragments. A higher proportion of polymorphic bands were observed using ISSR (85%) and semi-random (90%) primers than RAPD (69%) method. The combination of any two semi-random markers allowed the identification of all 20 bean genotypes. In contrast ISSR (except for primer (CAC)₃GC) and RAPD markers appeared to be less informative as more than two markers were necessary to achieve the same diagnostic level. Moreover, 7 ISSR, 2 RAPD and 8 semi-random exclusive bands were identified as putative population-specific markers. Semi-random and ISSR derived dendrograms showed similar tendencies in terms of genetic relatedness, whereas clustering of genotypes within groups was not similar when compared with the RAPD technique. Despite the different ability to resolve genetic variation among the investigated landraces, two major clusters with less than 60% (ISSR) and 40% (RAPD and semi-random) genetic similarity were formed with all three marker systems. The two groups were correlated with the phaseolin patterns and seed size of the landraces. The analysis showed that the cultivar ȁ8Lingua di Fuocoȁ9 and most of the landraces (13 out of 16) collected in Italy belong to the Andean gene pool, whereas only the three populations from Pratomagno belong to the Middle American gene pool.     

Characterization of Vitis germplasm using random amplified polymorphic DNA markers

Summary An analysis of the amplification fragments polymorphism of DNA coming from different accessions of germplasm belonging to species and cultivars of the genus Vitis, was carried out using 40 primer decamers of arbitrary sequence. The RAPD profiles showed a great intraspecific diversity. In many cases a single primer produced a unique pattern for each species. A phylogram tree based upon presence/absence data of the principal DNA bands divided the species according to their geographical origins. The intraspecific polymorphism of DNA fragments was not sufficient for an unambiguous identification of Vitis vinifera cultivars but the RAPD profiles turned out to be highly reproducible. The high capacity of this technique to generate DNA markers offers a new possibility for the study of the genetic relationships in the genus Vitis.Abbreviations PCR Polymerase chain reaction - RAPD Random amplified polymorphic DNA     

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.     

RAPD and ISSR molecular markers in <Emphasis Type="Italic">Olea europaea</Emphasis> L.: Genetic variability and molecular cultivar identification

Thirty Portuguese and eight foreign olive (Olea europaea L.) cultivars were screened using Random Amplified Polymorphic DNA (RAPD) and Inter-Simple Sequence Repeat (ISSR) markers. Twenty RAPD primers amplified 301 reproducible bands of which 262 were polymorphic; and 17 ISSR primers amplified 204 bands of which 180 were polymorphic. The percentage of polymorphic bands detected by ISSR and RAPD was similar (88 and 87%, respectively). The genetic variability observed was similar in the Portuguese and foreign olive cultivars. Seven ISSR and 12 RAPD primers were able to distinguish individually all 38 olive cultivars. Twenty specific molecular markers are now available to be converted into Sequence Characterised Amplified Region (SCAR) markers. Relationships among Portuguese and foreign cultivars is discussed.     

Genetic diversity and relationships among safflower (<Emphasis Type="Italic">Carthamus tinctorius</Emphasis> L.) analyzed by inter-simple sequence repeats (ISSRs)

Genetic diversity and relationships among 48 safflower accessions were evaluated using 22 inter-simple sequence repeats (ISSR) primers. A total of 429 bands were amplified, and 355 bands (about 82.7%) were polymorphic. Five to forty-one polymorphic bands could be amplified by each primer, with an average of 16.1 polymorphic bands per primer. The results showed that the polymorphism of the safflower germplasm was higher at the DNA level. All the 48 accessions could be distinguished by ISSR markers and were divided into 9 groups based on ISSR GS by using UPGMA method. The genetic relationships among the accessions from different continents were closer. Comparatively, the genetic diversity of the accessions originated from Asia was higher, from Europe assembled. The results also showed that the genetic variation of accessions from Indian and Middle Eastern safflower diversity centers were relatively higher. ISSR is an effective and promising marker system for detecting genetic diversity among safflower and give some useful information on its phylogenic relationships.     

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.     

Genetic Diversity among Wild Common Beans from Northwestern Argentina Based on Morpho-agronomic and RAPD Data

The genetic diversity among 10 wild populations of common bean Phaseolus vulgaris var. aborigineus was analyzed by means of RAPD markers and morpho-agronomic data. The study was performed on populations collected from different sites located in the provinces of Jujuy, Salta and Tucumán in northwestern Argentina. Ten quantitative traits and 33 random primers were scored. Clustering based on morpho-agronomic traits and RAPD markers generated similar phenograms that grouped bean populations based on their site of collection. The levels of diversity observed among populations were low suggesting they have a common ancestor. The levels of diversity shown by morpho-agronomic traits were higher compared to those of molecular markers, most probably due to the effect of the environment. Furthermore, a 480-bp DNA band identified a group of wild populations collected from similar sites. Breeding strategies need to exploit this diversity to broaden the genetic base of commercial beans to develop high yield cultivars.     

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.     

Assessing genetic diversity of sweet potato (Ipomoea batatas (L.) Lam.) cultivars from tropical America using AFLP

The sweet potato genebank at the International Potato Center (CIP) maintains 5,526 cultivated I. batatas accessions from 57 countries. Knowledge of the genetic structure in this collection is essential for rational germplasm conservation and utilization. Sixty-nine sweet potato cultivars from 4 geographical regions (including 13 countries) of Latin America were randomly sampled and fingerprinted using AFLP markers. A total of 210 polymorphic and clearly scorable fragments were generated. A geographic pattern of diversity distribution was revealed by mean similarity, multidimensional scaling (MDS), and analysis of molecular variance (AMOVA). The highest genetic diversity was found in Central America, whereas the lowest was in Peru-Ecuador. The within-region variation was the major source of molecular variance. The between-regions variation, although it only explains 10.0% of the total diversity, is statistically significant. Cultivars from Peru-Ecuador, with the lowest level of within region diversity, made the most significant contribution to the between region differentiation. These results support the hypothesis that Central America is the primary center of diversity and most likely the center of origin of sweet potato. Peru-Ecuador should be considered as a secondary center of sweet potato diversity.     

Inter Simple Sequence Repeat (ISSR) Analysis of Diploid Coffee Species and Cultivated Coffea arabica L. from Tanzania

Inter simple sequence repeat (ISSR) markers were used to evaluate levels of genetic similarity among Coffea arabica L. accessions from Tanzania and to estimate levels of genetic similarities in C. arabica and diploid coffee species. The six ISSR primers used generated a total of 82 fragments and the dissimilarity values ranged from 0.21 to 1. Mean dissimilarity values between provenances (0.56–0.85) were higher than within provenances (0.37–0.68). Cluster analysis based on Nei’s genetic distances showed C. arabica provenances grouping based on geographical origin. Two major clusters were formed that constituted of provenances from Kilimanjaro and Arusha in one sub-cluster; Tanga and Morogoro in the other; the second cluster had Mbeya provenances and diploid species, respectively. The implication is that Mbeya provenances are different from the rest of Tanzanian C. arabica. A principal coordinate analysis (PCA), whose first three coordinates explained 43% of the variation, showed similar groupings as in the cluster analysis. A separate cluster analysis of diploid species showed a distinct separation of the three species used. ISSR data gave results similar to previous findings from random amplified polymorphic DNA(RAPD) analysis. The results also confirm the limited diversity present in cultivated C. arabica in Tanzania     

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.     

Genetic diversity of the complex Paspalum notatum Flügge (Paniceae: Panicoideae)

The genus Paspalum L. comprises approximately 400 species worldwide and about 220 in Brazil. Paspalum is ecologically and economically important, and has been very useful as pasture and P. notatum Flügge (bahiagrass) is a valuable forage grass in the subtropics. This species consists of several sexual (diploid) and apomictic (tetraploid, ocasionally tri and pentaploids) biotypes. In this work, inter simple sequence repeats (ISSR) markers were used to assess the genetic variability of a bahiagrass (P. notatum) collection. Vegetative tissues of 95 bahiagrass accessions were obtained from various locations in South America (Brazil, Argentina and Uruguay). A total of 91 reproducible ISSR fragments were observed and 89 fragments (97.5% of the total observed) were polymorphic. Cluster analyses (UPGMA) were performed from the ISSR data set and the results illustrate the genetic relationships among the 95 accessions of P. notatum. A comparison among molecular, morphological and ploidy levels data were done. ISSR markers were effective in distinguishing the genotypes analyzed, and a wide variability was observed for this species. These results add new information regarding the genetic diversity in P. notatum, thus contributing toward the biological knowledge of this species, and providing with subsides for future plant breeding and conservation programs.     

Diversity of 21 taro (Colocasia esculenta (L.) Schott) accessions of Andaman Islands

The genetic diversity of taro (Colocasia esculenta (L.) Schott.) accessions growing naturally in Andaman Islands was analysed using morphological and DNA markers. Twenty one representative samples of C. esculenta from different parts of Islands in addition to three commercial varieties as reference genotypes were used in study. About 63% phenotypic variation was observed in C. esculenta A total number of 491 amplified fragments were obtained of which 347 showed polymorphic banding patterns. The accessions were grouped into two major clusters with both RAPD and ISSR markers with 56 and 57% diversity, respectively. The reference genotypes were grouped into one group and island population in other cluster. Both marker systems divided population into two sub clusters and showed correlation with morphological parameters. The diversity pattern observed in present study showed rich genetic diversity of C. esculenta in Andaman Islands provided simple strategy for reducing repeatability of taro germplasm in gene banks. The study also suggested pre-evaluation of germplasm using molecular and morphological markers to enhance efficiency of exploration trips.     

A preliminary study of the genetic diversity of Bolivian oca (<Emphasis Type="Italic">Oxalis tuberosa</Emphasis> Mol.) varieties maintained<Emphasis Type="Italic"> in situ</Emphasis> and<Emphasis Type="Italic"> ex situ</Emphasis> through the utilization of ISSR molecular markers

ISSR molecular markers have been used to investigate genetic diversity of oca (Oxalis tuberosa Mol.), an Andean neglected tuber crop species. Sampling procedure allowed a preliminary study of the genetic diversity at the intra- and intervarietal levels. Twenty tuber lots conserved in situ in the microcentre of Candelaria and ex situ in the Toralapa Centre (Bolivia) were identified. Four ISSR primers amplified a total of 25 fragments of which 17 (68%) were polymorphic. These experiments show that the structure of oca varieties is mainly based upon vernacular names with a greater differentiation among tuber lots than within them, supporting agromorphological data. ISSR technique enlightened the existence of heterogeneous varieties in oca and divergence between in situ and ex situ conservation strategies. These observations are potentially linked to the different ways of management of tubers in these two conservation systems.     

Genetic variation among populations of wild safflower, <Emphasis Type="Italic">Carthamus oxyacanthus</Emphasis> analyzed by agro-morphological traits and ISSR markers

Wild species of safflower, Carthamus oxyacanthus Bieb., is highly crossable with cultivated species, C. tinctorius L. and could be directly exploited in broadening safflower gene pool and improving the crop for biotic and abiotic stress environments. In this study, genetic diversity among accessions of C. oxyacanthus and their relationships with cultivated safflower were evaluated using agro-morphological traits and polymorphic inter-simple sequence repeats (ISSR) markers. Significant variation was observed among accessions particularly for seeds per capitulum, seed yield per plant, harvest index and capitula per plant. Cluster analysis based on agro-morphological traits classified the wild accessions in two groups according to their geographical regions, and separated them from the cultivated genotypes. ISSR marker also revealed a high genetic variation among the accessions, and cluster analysis based on this marker divided genotypes into four groups, with cultivated ones in a separate clade. Genetic variation observed among the wild safflower germplasm at the DNA level was higher than the agro-morphological traits, indicating that ISSR is an effective marker system for detecting diversity among safflower genotypes and their genetic relationships. Accessions of C. oxyacanthus with high genetic relationship to cultivated species could be used for interspecific hybridization in breeding programs of safflower.     

Analysis of genetic diversity among European and Asian fig varieties (<Emphasis Type="Italic">Ficus carica</Emphasis> L.) using ISSR,RAPD, and SSR markers

Nineteen fig varieties and lines from Europe and Asia have been fingerprinted by ISSR, RAPD, and SSR markers, respectively, using 13, 19, and 13 primer combinations. All primers produced 258 loci, with the highest number of loci (119) generated by RAPD (R _p: 48.42). Clustering analysis was applied to the three marker datasets to elucidate the genetic structure and relationships among these varieties. Mean genetic similarities were 0.787, 0.717, and 0.749, respectively, as determined using ISSR, RAPD, and SSR. Each marker system produced incompletely separated clusters, although a weak binding group based on race type appeared in the combined dataset. Comparisons of coefficients revealed no correlation between different similarity matrices; congruence was observed between similarity matrices and co-phenetic matrices in all markers. Analysis of molecular variance (AMOVA) showed that most of the total polymorphism was attributable to within-group variance (ISSRs + RAPDs, 97.41%; SSRs, 90.18%). These results suggest that the genetic diversity of this fig population is low and that multiple marker utilization is critical to estimate the relatedness of figs at the variety level. Additionally, it was presumed that ‘Houraihi’, the oldest variety in Japan, was disseminated independently of other foreign varieties in the 17th century or before then.