Inter-simple sequence repeat (ISSR) and RAPD variation among wild barley (Hordeum. vulgare subsp. spontaneum) populations from west Turkey

Inter-Simple Sequence Repeat (ISSR) and Randomly Amplified Polymorphic DNA (RAPD) markers were used to analyze genetic distance among H. vulgare subsp. spontaneum populations from west Turkey. Fifty-five RAPD and 10 ISSR primers were used to detect variation among sample. A total of 55 polymorphic loci were found using 65 primers. Two distinct cluster groups were clearly established among populations. The minimum variation was detected between Pinarbasi and Bornova (GD = 0.192) populations and the maximum was found between Icmeler and Aydin populations (GD = 0.926). As two dominant markers, RAPD and ISSRs are effective and promising marker systems for detecting genetic variation.     

Genetic variation in cucumber (Cucumis sativus L.) as assessed by random amplified polymorphic DNA1

Isozyme and restriction fragment length polymorphisms (RFLPs) have been applied to studies of genetic relationships and germplasm management in cucumber (Cucumis sativus L.). However, isozymes identify relatively few polymorphisms, and RFLPs are technically complex, expensive, and not compatible for the high through-put required for rigorous assessment of this narrow-based germplasm. Since random amplified polymorphic DNA (RAPD) markers do not manifest such shortcomings, a study was conducted in cucumber to examine genetic relationships in diverse germplasm, assess the usefulness of RAPD markers in distinguishing elite accessions, and compare the relative effectiveness of RAPD markers to that of isozyme and RFLP markers. One hundred and eighteen C. sativus accessions were analyzed using variation at 71 RAPD loci (44 mapped and 27 unmapped). Genetic distances among accessions were estimated using the simple matching coefficient complement, and analyzed using multi-dimensional scaling. Each accession had a unique marker profile, indicating that RAPD analysis was useful in genotypic differentiation. Germplasm grouping patterns were consistent with individual accession origins, theoretical dispersal routes and discriminating morphological characters (i.e., sex expression and fruit length to diameter ratio). Although elite accessions were discriminated by RAPD profiling, their genetic distances were relatively small (between 0.01 and 0.58), indicating limited genetic diversity in this germplasm array. Assessment of a subset of the germplasm array using RAPDs resulted in genetic distance measurements more similar to published genetic distance estimates by RFLP markers (Spearman rank correlation, r_s = 0.7–0.8) than estimates by isozyme markers (r_s = 0.4). Data indicate that RAPD markers have utility for analysis of genetic diversity and germplasm management in cucumber.     

Genetic Diversity Among Brazilian Cultivars and Landraces of Tomato Lycopersicon Esculentum Mill. Revealed by RAPD Markers

Random amplified polymorphic DNA markers (RAPD) were used to estimate the variability of 35 tomato accessions (Lycopersicon esculentum Mill.). A total of 257 reproducibly scorable bands were obtained from 20 primers, 78.6% of which were polymorphic. The percentage distribution of RAPD markers shows a bimodal distribution, and the frequency of rare alleles is similar in commercial and landrace accessions. Genetic distances among accessions were calculated and a dendrogram showing the genetic relationships among them was constructed allowing for the separation of four groups. Twenty out of 23 Brazilian landraces fell within one group, whereas commercial cultivars were distributed in the four groups. AMOVA analysis of RAPD data showed that, despite the high within Brazilian landraces and commercial cultivars variation, these two groups are significantly different, indicating that landraces can be a source of variation for breeding programs.     

Assessment of genetic variation in a working collection of Vigna vexillata (L.) A. Rich. by isozyme and RAPD analyses

Genetic variation based on isozyme and RAPD analyses was investigated in 47 and 34 accessions respectively of Vigna vexillata from different geographical origins and belonging to three botanical varieties. A total of 9 enzyme systems were studied, accounting for 14 putative loci, 8 of which were polymorphic. The analysis of genetic diversity revealed a low level of within accession variation (H_S=0.013), while between accession diversity (D_ST) was 0.120. Coefficient of gene differentiation (G_ST) was 0.905, indicating that most variation was among accessions. Nei's genetic distances were calculated on the basis of allelic frequencies and a UPGMA dendrogram was constructed. Twenty arbitrary 10-mer oligonucleotides were used in RAPD analysis. Amplification profiles disclosed a higher level of polymorphism than isozymes. Based on amplification patterns, the similarity index of Jaccard was calculated and a dendrogram constructed on the basis of the similarity matrix. The final clustering based on RAPD data was similar to the one obtained using isozyme allelic frequencies. The classification in botanical varieties did not reflect the allelic constitution of the different samples. On the other hand, referring to geographical origin, most accessions from Africa and from Latin America were distributed respectively in two distinct clusters in the dendrogram. This grouping might also reflect the differences observed in the germination behaviour of V. vexillata from the two continents.     

Comparison of isozyme and random amplified polymorphic DNA data for determining intraspecific variation in Cucumis

Variation at isozyme and random amplified polymorphic DNA (RAPD) loci in eight cucumber and seven melon cultivars, breeding lines, and plant introductions were used to determine the utility of these markers for assessing genetic variation among populations of each species. Although dendrograms derived from cluster analyses using species' variation at marker loci were dissimilar, these disparities were consistent with differences in the pedigrees and/or other information (e.g., morphological) known about each accession and species. Empirical estimations of variances associated with each marker type in the cucumber and melon accessions examined indicate that, per band, lower coefficients of variation can be attained in the estimation of genetic difference when using RAPDs compared to isozymes. The disparity between the marker analyses made may be related to the amount of genome coverage characteristic of a particular marker system in a species and its efficiency in sampling variation in a population.     

Genetic variability in Turkmen populations of Pistacia vera L.

Seedlings of Pistacia vera L. developed from seeds of two separate populations in Turkmenistan, Kepele and Agachli, were evaluated for their growth potential and genetic polymorphism. Plant growth rate as well as random amplified polymorphic DNA (RAPD) analysis showed distinct differences between the two populations. In plant height growth rate, 17 Agachli accessions were 1.3 times higher on average than that of 10 accessions of Kepele (significant at p = 0.046) and 1.2 times higher for trunk diameter growth rate (p = 0.062). Cluster analysis divided most accessions into two main genetic groups according to their geographic origin. The Agachli group was further divided into two subgroups. One Kepele accession (K4), was genetically different from the rest and clustered on a separate outgroup. Two Agachli accessions (A12 and A17) were outside the two main populations clusters. Accessions K9 and K10 from Kepele were exceptional and were clustered in each of the two Agachli's subgroups, indicating a close genetic relationship between the two populations. In addition, high similarity values (0.58–1.00) and small genetic distances reflect plausible gene flow between Kepele and Agachli, which are 100 Km apart. Mantel test revealed significant relationship between the RAPD and the morphological traits matrices, pointing to the genetic basis for the measured differences in the growth rate. Growing the accessions on the same plot, under similar conditions enabled the evaluation of genotypic differences. The combination of morphological traits and molecular markers will further assist in preservation of genetic variability and cultivation of useful genotypes of P. vera L.     

Relationships between an Italian Strawberry Ecotype and its Ancestor using RAPD Markers

Random-amplified polymorphic DNA (RAPD) markers were used to evaluate genetic variability among populations of an Italian strawberry ecotype, and to determinate genetic relationships between genotypes and their putative ancestor. A total of 65 selections and one cultivar ‘Madame Moutot’ (MM), were analysed to evaluate genetic variability present in Etna mountain area and to confirm as MM was one of the cultivars that originated the ecotype. A total of 222 RAPD markers was obtained using 16 decamer primers and 6 longer primers, 90.8% of the markers obtained by selected primers resulted polymorphic at least within analysed genotypes. RAPDs were used to calculate genetic similarity coefficients and to generate dendrograms representing genetic relationships among genotypes analysed. Cluster analysis displays as RAPD polymorphisms were able to characterize the genotype variability among closely related groups. The data show as MM could be considered the ancestral genotypes introduced in that area. The results obtained confirm that RAPD markers could be used as reliable markers to perform phylogenetic studies in Fragaria×ananassa Duch. ex Rozier. Giuseppe Bertino and Piero Spada - Coauthor involved in genotype selection and field management     

Levels and distribution of allozyme and RAPD variation in populations of Elymus fibrosus (Schrenk) Tzvel. (Poaceae)

To study the magnitude and nature of genetic variation in E. fibrosus, the levels and distribution of allozyme and RAPD variations were investigated in populations collected from Finland and Russia. The results obtained from the allozyme and RAPD studies were compared to each other in 10 of the populations. The allozyme analysis showed that 6 of 12 presumed loci (50%) were polymorphic within the species, while the mean number of polymorphic loci within populations was 4.8%. The mean number of allele per locus for the species was 1.5 and 1.05 across the populations. Genetic diversity at the species level was low (H _es = 0.025), and the mean population genetic diversity was even lower (H _ep = 0.007). Both these values were much lower than the average for other Elymus and self-fertilising species. The largest proportion of the total allozyme diversity was found among, rather than within the populations (G _ST = 0.70). The allozyme genetic distances between the populations did not reflect geographic distances. Cluster and principal coordinates analyses revealed the same allozyme relationship patterns among the populations. A comparison of allozyme and RAPD variation in 10 of the populations showed differences in the amount of genetic variation. The RAPD analysis revealed higher levels of variation (A _p = 1.19, P _p = 20.3 and H _ep = 0.09) than the allozyme one) A _p = 1.06, P _p = 5.8 and H _ep = 0.008). For both markers, the largest proportion of the total gene diversity was found among the populations studied (G _st = 0.63 for RAPDs and G _st = 0.65 for allozyme). In contrast to the allozyme analysis, the RAPD based genetic distances did reflect geographic distances. The cluster and principal coordinates analyses showed different grouping of populations for each data set. There was a positive, but not significant, correlation (r = 0.41) between the genetic distance matrices resulting from these markers. Regional comparison revealed that the Finnish populations had a higher diversity than the Russian ones. Generally, this study indicates that E. fibrosus contains low genetic variation in its populations. The results are discussed in the context of conservation of the species.     

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 diversity within a range of cultivars and landraces of flax (Linum usitatissimum L.) as revealed by RAPDs

Analysis of the extent and distribution of genetic diversity incrop plants is essential for optimizing sampling and breedingstrategies. We used random amplified polymorphic DNA (RAPD)markers to assess genetic diversity and relationships in 22 Canadiancultivars, 29 selected world cultivars and 10 landraces of flax(Linum usitatissimum L.). RAPDvariation was generally low and more variation was detected among,than within, the investigated flax accessions. Based on 53 variableRAPD loci observed for the 61 accessions, the landraces had a lowerproportion of fixed recessive RAPD loci (0.427) (i.e.,more genetic variation) than all of the flax cultivars examined(0.492). The linseed cultivars had a lower proportion ofrecessive loci (0.469) than the fiber flax cultivars(0.529). Canadian linseed cultivars had a lower proportionof recessive loci (0.465) than the selected world flaxcultivars (0.512). A trend was also observed that the rateof loss in genetic variation in Canadian flax breeding programs overthe last fifty years was approximately two variable loci per 100 lociper 10 years. Clustering analyses based on similarity estimatesshowed that the fiber cultivars were more related (or similar toeach other) and were classified as a homogeneous group. All ofthe linseed cultivars were clustered in diverse groups with the ninelandrace accessions. Implications of these findings for flax breedingand germplasm management are discussed.     

Comparative analysis of genetic diversity using molecular and morphometric markers in Andrographis paniculata (Burm. f.) Nees

Andrographis paniculata is a medicinal plant of immense therapeutic value. The present study was aimed to elucidate its genetic diversity based on morphochemical and RAPD markers from 53 accessions belonging to 5 eco-geographic regions. Analysis of variance and D ² statistics revealed significant differences in all the metric traits and sufficient inter-cluster distances indicating considerable diversity among the accessions. The complementary approach of RAPD was used to evaluate the genetic dissimilarities among all the accessions using 6 highly polymorphic primers. The average proportion of polymorphic loci across primers was 96.28%. The molecular genetic diversity based on Shannon index per primer averaged 5.585 with values ranging from 3.08 to 8.70 indicating towards wide genetic base. RAPD based UPGMA and D ² cluster analysis also revealed that various accessions available in different eco-geographic regions might have originated from native places of wild abundance. Similarity matrices were generated for molecular markers and morphometric data to determine the degree of congruence between the two. A highly significant but low correlation (r = 0.547, P < 0.001) was obtained thus implying the correspondence between the two. The species is hermaphroditic and a habitual inbreeder. The present study yielded a typical triangular congruence between its breeding system, morphometric traits and RAPD markers thus elucidating the usefulness of complementary approaches to make diversity analysis more explanatory and purposeful for optimum genetic amelioration and effective conservation of its genotypic variability.     

Differences in the apportionment of molecular and morphological variation in North American strawberry and the consequences for genetic resource management

Variation for 24 morphological traits measured in a greenhouse environment and 36 randomly amplified polymorphic DNA (RAPD) markers was assessed among 318 wild octoploid strawberry (Fragariaspp.) genotypes from diverse habitats across the northern USA. RAPD marker frequencies and certain leaf and flower morphology traits (petiole color, leaf mass/area ratio, leaflet length and width, flower and receptacle diameter, petal width, flowers/inflorescence) were significantly different between the F. chiloensis-platypetala and F. virginiana-glauca species complexes. The proportion of variation accounted for by provenance effects was lower for the RAPD markers than for most morphological traits, especially in the F. virginiana-glauca species complex. Morphological traits of potential adaptive importance group the collection into provenances within each species-complex, and reflect the significant habitat and geographic differences across the region from which the germplasm was collected. Variation among populations within provenances was low for the molecular and most morphological traits, with a much larger amount of variability among plants within populations. Most of the variation for the presumably more selectively-neutral RAPD data was among plants within populations and populations within provenances rather than among the provenances that were recognized based on morphological traits, especially in the F. virginiana-glauca complex. Patterns of diversity for morphological traits must be considered, along with more selectively-neutral molecular characters such as RAPDs, to formulate effective sampling strategies and to properly estimate the quantity and apportionment of diversity within this germplasm.     

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.     

Genetic variability of Coffea arabica L. accessions from Ethiopia evaluated with RAPDs

The genetic diversity of 50 wild and semi-wild accessions of the Coffea arabica L. germplasm collection, gathered by the FAO and ORSTOM missions to Ethiopia, and maintained in Colombia by CENICAFE, was evaluated with RAPD markers. The evaluation was carried out in two phases: In phase one, the polymorphism of 8 Ethiopian accessions of different geographic origin, plus the cultivated variety 'Caturra' was assessed with the RAPD technique with forty-two 10-mer oligonucleotides. In phase two, 51 accessions were assessed with a set of 5 polymorphic primers that reproduced, with a correlation of 95%, the groups generated by the 24 polymorphic primers found in phase one. Principal Coordinate Analysis of molecular data revealed that a closely related group consisting of 86% of the Ethiopian C. arabica accessions evaluated are significantly different from the Caturra variety and could be used in a genetic breeding initiative to increase the variability of cultivated varieties. The results also indicate that a larger polymorphism is present in the Colombian replica of FAO Ethiopian coffee germplasm collection than previously reported.     

Microsatellites and RAPD Markers to Study Genetic Relationships Among Cowpea Breeding Lines and Local Varieties in Senegal

Genetic diversity in local cowpea varieties and breeding lines from Senegal were studied using random amplified polymorphic DNA (RAPD) and microsatellite (SSR) techniques. Among the 61 RAPD primers used, twelve show polymorphism. Fifteen of the 30 microsatellite primer pairs were polymorphic, detecting one to nine alleles per locus. The RAPD and SSR data were analyzed both separately and in combination to assess relationships among genetic lines. Although RAPD provided information on levels of genetic diversity, microsatellite markers are most effective in determining the relationship among cowpea accessions and varieties. The SSR results support the genetic diversification of cowpea in Senegal and underscore their potential in elucidating patterns of germplasm diversity of cowpea in Senegal. An erratum to this article is available at .     

Evaluation of the discriminance capacity of RAPD, isoenzymes and morphologic markers in apple (Malus x domestica Borkh.) and the congruence among classifications

Twenty-one apple accessions were characterised and classified taxonomically with randomly amplified polymorphic DNA (RAPD). The results were compared with morphological and isoenzymatic classifications of the same material by calculating the congruence between similarity matrices. The results confirmed the greater discrimination capacity of RAPD compared with isoenzymes, although the taxonomic clusters obtained with both types of markers were congruent. The genetic relationships calculated using RAPD and isoenzymes showed little congruence with the morphological relationships among accessions. Isoenzymatic markers are useful in germplasm banks to evaluate the diversity of newly introduced accessions. The use of RAPD markers are especially beneficial to discriminate between material that is genetically similar, to evaluate genetic variability within a collection and to choose the components of the core collection.     

Genetic diversity in Sorghum bicolor(L.) Moench accessions from different ecogeographical regions in Malawi assessed with RAPDs

Genetic variation within and among several Sorghum populations from different agroecological zones in Malawi were investigated using random amplified polymorphic markers (RAPDs). DNA samples from individual plants were analyzed using 35 oligonucleotides of random sequence. Twenty five of these primers allowed amplifications of random polymorphic (RAPD) loci. Overall, 52% of the scored loci were polymorphic. Every accession was genetically distinct. The analysis of molecular variance revealed that the within-region (among accessions) variations accounted for 96.43% of the total molecular variance. Observed variations in allelic frequency was not related to agroecological differences. The degree of band sharing was used to evaluate genetic distance between accessions and to construct a phylogenetic tree. Further analysis revealed that the sorghum accessions analyzed were genetically close despite considerable phenotypic diversity within and among them. It is suggested that all the sorghum landraces currently available in Malawi should be conserved both ex situ and in situ to maintain the current level of genetic diversity.     

Morphological and molecular diversity analysis among the Indian clones of <Emphasis Type="Italic">Sesuvium portulacastrum</Emphasis> L.

Sesuvium portulacastrum L. (seapurslane) is a halophyte used as pioneer species in sand dune fixation and stabilization of saline soil. Studies on the morphological and molecular diversity were carried out for the 14 clones of Sesuvium collected from the different coastal regions of India. Significant differences were observed for morphological traits viz., length, width, diameter and area of leaf, internodal distance and stem diameter for different clones when compared with the clone from Gujarat state (GJ1). A UPGMA dendrogram for morphological traits based on the Pearson’s similarity coefficient clustered the clones into three groups considering 80% polymorphism as criteria. Molecular diversity among the clones was studied using Randomly Amplified Polymorphic DNA (RAPD), Internal Transcribed Spacer (ITS) and markers specific to Ac homologous region. Of the total 749 RAPD loci amplified with 70 random primers, 294 were polymorphic with 39.25% diversity. A phylogenetic tree constructed with UPGMA and SHAN, grouped the clones into three major clades based on RAPD data. The molecular diversity studied with ITS and markers specific to Ac homologous region revealed 37.50% and 66.66% polymorphism and clustered the clones into three and four clades, respectively. The genetic diversity analysis revealed wide variations among the S. portulacastrum clones, reflecting a high level of diversity within the species which might be due to anthropogenic impact and geographic environmental conditions. Further, the various clones from the different eco-geographic coastal localities might have originated from native places of wild abundance. To the best of our knowledge, this is the first attempt to evaluate both morphological and genetic diversity among the Sesuvium clones collected from the distant habitats of the coastal regions of the India.     

Assessing the Genetic Diversity in the International Cocoa Genebank,Trinidad (ICG,T) using Isozyme Electrophoresis and RAPD

Random amplified polymorphic DNA (RAPD) and isozyme electrophoresis (IE) techniques were used to estimate the level of genetic diversity in a sample of cacao germplasm existing at the International Cocoa Genebank, Trinidad. Twenty-six cocoa populations represented by 459 cocoa genotypes were analysed using IE and 22 populations represented by 353 cocoa genotypes were analysed using RAPD. Despite few differences in the classification of the populations, both techniques revealed three major groups: the indigenous trees, the cultivated Trinitario and the cultivated trees from Ecuador. Two-thirds of the partitioned diversity were found within populations and one-third between the populations, with both techniques.     

Genetic Relationships and Variation among Biotypes of Dallisgrass (Paspalum dilatatum Poir.) and Related Species Using Random Amplified Polymorphic DNA Markers

The genus Paspalum L. consists of more than 400 species. Around twenty-five informal groups of species are recognized in Paspalum and the Dilatata group is of special interest because its members are excellent potential forage grasses. Seventy-five germplasm accessions, representing 15 taxa, were analyzed using randomly amplified polymorphic DNA (RAPD). Polymorphisms were observed with twenty-two primers in the Dilatata group and 16 of those were analyzed. Four hundred and four different RAPD fragments were generated, resulting in an average of 25.2 bands per primer. Among the 404 markers analyzed, 48 (11.88%) were exclusive for the P. dilatatum Poir. biotypes, 31 (7.67%) were exclusive to taxa belonging to other groups included in this study, 28 markers (6.93%) were diagnosed for other species of the Dilatata group and 16 (3.96%), for natural hybrids. Extensive RAPD variation was found among the species studied. Inter- and intra-taxonomic polymorphisms were detected. A dendrogram based on the RAPD data shows some clusters corresponding to the same taxa. However, the biotypes of P. dilatatum do not form a cluster. The present work confirms that the RAPD technique can be used to determine genetic relationships between the taxa belonging to the Dilatata group.