Genetic relationships among melon breeding lines revealed by RAPD markers and agronomic traits

 RAPD markers and agronomic traits were used to determine the genetic relationships among 32 breeding lines of melon belonging to seven varietal types. Most of the breeding lines were Galia and Piel de Sapo genotypes, which are currently being used in breeding programmes to develop new hybrid combinations. A total of 115 polymorphic reliable bands from 43 primers and 24 agronomic traits were scored for genetic distance calculations and cluster analysis. A high concordance between RAPDs and agronomic traits was observed when genetic relationships among lines were assessed. In addition, RAPD data were highly correlated with the pedigree information already known for the lines and revealed the existence of two clusters for each varietal type that comprised the lines sharing similar agronomic features. These groupings were consistent with the development of breeding programmes trying to generate two separate sets of parental lines for hybrid production. Nevertheless, the performance of certain hybrids indicated that RAPDs were more suitable markers than agronomic traits in predicting genetic distance among the breeding lines analysed. The employment of RAPDs as molecular markers both in germplasm management and improvement, as well as in the selection of parental lines for the development of new hybrid combinations, is discussed. Received: 25 July 1997 / Accepted: 6 October 1997     

Genetic diversity and genetic relationships of Chukrasia spp. (Meliaceae) as revealed by inter simple sequence repeat (ISSR) markers

Key message

ISSR characterization of Chukrasia populations from the natural range revealed two distinct groups of populations consonant with morphological differentiation. Results suggest the current taxonomic classification of the genus should be reviewed.

Abstract

There are different views as to whether the genus Chukrasia (Meliaceae) consists of one species, C. tabularis, or two species C. tabularis and C. velutina. Despite a clear pattern of variation in many morphological characteristics such as leaves and bark, some authors regard the latter merely an ecotype of the former in seasonal forest. In the present study, we used ISSR markers to determine the genetic diversity and population structure among 23 Chukrasia subpopulations from across the natural range in Asia. Molecular analysis clearly differentiated two distinct groups of subpopulations, corresponding to the putative species, as well as well-defined subpopulations corresponding to geographic regions within the two groups. The molecular results are in concordance with morphological differentiation and corresponded to the two recognized taxa. The present study suggests that current taxonomic classification of the genus Chukrasia should be reviewed.     

Genetic diversity and relationships in mulberry (genusMorus) as revealed by RAPD and ISSR marker assays

Background  

The genus Morus, known as mulberry, is a dioecious and cross-pollinating plant that is the sole food for the domesticated silkworm, Bombyx mori. Traditional methods using morphological traits for classification are largely unsuccessful in establishing the diversity and relationships among different mulberry species because of environmental influence on traits of interest. As a more robust alternative, PCR based marker assays including RAPD and ISSR were employed to study the genetic diversity and interrelationships among twelve domesticated and three wild mulberry species.     

Genetic diversity in Elymus caninus as revealed by isozyme, RAPD, and microsatellite markers.

Genetic diversity of 33 Elymus caninus accessions was investigated using isozyme, RAPD, and microsatellite markers. The three assays differed in the amount of polymorphism detected. Microsatellites detected the highest polymorphism. Six microsatellite primer pairs generated a total of 74 polymorphic bands (alleles), with an average of 15.7 bands per primer pair. Three genetic similarity matrices were estimated based on band presence or absence. Genetic diversity trees (dendrograms) were derived from each marker technique, and compared using Mantel's test. The correlation coefficients were 0.204, 0.267, and 0.164 between isozyme and RAPD distance matrices, RAPD and microsatellite distance matrices, and between isozyme and microsatellite distance matrices, respectively. The three methodologies gave differing views of the amount of variation present but all showed a high level of genetic variation in E. caninus. The following points may be drawn from this study whether based on RAPD, microsatellite, or isozyme data: (i) The Icelandic populations are consistently revealed by the three dendrograms. The congruence of the discrimination of this accession group by RAPD, microsatellite, and isozyme markers suggests that geographic isolation strongly influenced the evolution of the populations; (ii) The degree of genetic variation within accessions was notably great; and (iii) The DNA-based markers will be the more useful ones in detecting genetic diversity in closely related accessions. In addition, a dendrogram, which took into account all fragments produced by isozymes, RAPDs, and microsatellites, reflected better the relationships than did dendrograms based on only one type of marker.     

Genetic diversity among Turkish local grape accessions (Vitis vinifera L.) using RAPD markers

To estimate genetic relationships among 46 local grape cultivars, RAPD analysis was performed with 25 decamer primers selected from a total of 60 primers. Genetic relationships among these cultivars were determined by calculating similarity indexes, from which a dendogram was derived. There was high genetic variation among the cultivars, with values of genetic diversity ranging from 0.553 to 0.952 using the Jaccard coefficient. UPGMA analysis of a distance matrix produced a dendogram with six clusters. The relatively high genetic similarity ratios observed for the cultivars was also reflected in the dendogram. In general, no relationship was encountered between the genetic similarity ratios of the cultivars and the results of previous ampelographic analyses.     

Genetic relationships among ten endod types as revealed by a combination of morphological,RAPD and AFLP markers

The genetic relationships among ten types of endod (Phytolacca dodecandra) cultivated by the Institute of Pathobiology of the Addis Ababa University to combat the disease bilharzia in Ethiopia were studied using morphology and molecular markers. A total of 18 morphological characters, 194 amplified fragment length polymorphism (AFLP) and 42 random amplified polymorphic DNA (RAPD) markers were used to determine genetic proximity between types. Genetic distance and cluster analysis of the AFLP data revealed the lack of genetic difference between E47 and E48 but relatively wider genetic difference among the other endod types. Cluster and principal component analyses performed on the AFLP and RAPD markers demonstrated the presence of distinct separation of E56 but not that of E44 from the others. The AFLP and RAPD data, thcrefore, did not support the hypothesis that the superiority of E44 in agronomic traits and molluscicidal potency is linked to its distinct genetic difference from the other endod types. Matrices correspondence tests demonstrated the presence of greater correspondence between AFLP and RAPD data (r = 0.842) but not between the morphology and that of AFLP and RAPD. This indicates the correspondence more between the two DNA markers systems than either of them with morphological traits. The cophenetic correlation coefficients also revealed poor fit for morphology (r = 0.716), good fit for RAPD (r = 0.872) and very good fit for AFLP (r = 0.975), reflecting the hyper-variability and higher resolving power of AFLP.     

Phylogenetic relationships among Portuguese rye based on isozyme, RAPD and ISSR markers

The phylogenetic relationships of 10 rye landraces and cultivars from the north of Portugal and from Brazil were analysed using 20 isozyme loci, and a total of 511 PCR markers (342 ISSRs and 169 RAPDs). The isozymes were analysed in at least 100 plants of each population/cultivar and, therefore, we have data about intra and inter population/cultivar genetic variability. However, the analyses with ISSRs and RAPDs were obtained using a mix of 25 plants of each population. Therefore, each population/cultivar was reduced to one tube and we have no data about intra genetic variability. As expected in a cross pollinated crop we found genetic diversity and a larger variation within than among the populations using isozymes. Somewhat unexpectedly, however, we found that the breeding cultivars have the same level of heterozygosity as the landraces. The phylogenetic relationships obtained using isozymes among the landraces, synthetic cultivar and the cultivars from breeding programs do not reflect their origin. Moreover, the cultivar from Brazil is not separated from the remaining populations/cultivars studied. However, the data observed using RAPDs and ISSRs are in agreement with their known origin. The populations maintained by the farmers in the north of Portugal are grouped in a cluster in the phenogram and the C902591 (from Brazil), the Alv?o (synthetic variety) and Larouco (a hybrid between Montalegre and Brazil) are in a different cluster. The ISSRs and RAPDs provide a rapid method for the production of polymorphic markers, which appear to correspond to known pedigree information.     

Assessment of genetic diversity in Colletotrichum falcatum Went accessions based on RAPD and ISSR markers

Sugarcane is susceptible to red rot disease caused by phytopathogenic fungus Colletotrichum falcatum Went which ultimately affect the economy of farmers as well as sugar based industry. One of the various ways to control this devastating disease is to develop disease resistance sugarcane cultivar and this requires the complete understanding of genetic makeup of pathogen. Although South Gujarat is well known sugarcane cultivating area, less published data can be found about PCR-based genetic diversity in prevalent C. falcatum accessions. So, present investigation aims at finding molecular variation among the ten accessions of C. falcatum using RAPD and ISSR molecular markers. A total of 35 RAPD and 39 ISSR primers were screened across 10 C. falcatum accessions, of which 15 RAPD and 21 ISSR primers have showed consistent amplification. Statistics related to genetic variation were estimated using NTSYS-PC by means of Dice’s coefficient. The results revealed 80.6% and 68.07% polymorphism and similarity coefficient ranged from 0.43 to 0.91 and 0.73 to 0.93 in RPAD and ISSR analysis respectively. The dendrogram generated using RAPD, ISSR and combined RAPD-ISSR grouped accessions into different clusters which reveal considerable level molecular variation among the C. falcatum accessions. It is also evident from PCA plots that accessions are rather dispersed with tested marker systems indicating good genetic base. So, in nut shell, we found considerable genetic variation and relatedness within C. falcatum accessions collected from different areas of south Gujarat, India using RAPD and ISSR markers.     

Genetic diversity in a collection of Cucumis sativus L. assessed by RAPD and ISSR markers

The genetic diversity among 39 cucumber collections from Karnataka, India was assessed using 23 RAPD and 18 ISSR primers. A total of 309 bands were scored of which 147 (47.57 %) were polymorphic. The average number of bands per primer was 7.82 and an average number of polymorphic bands of 3.58 per primer. The primers UBC855 revealed the highest PIC (polymorphism information content) value of 0.49 followed by the primers UBC846, OPE13, OPC01 and OPR12 (0.48). The Jaccard’s similarity coefficients ranged from 0.36 to 0.84 and the first two principal components explained 53.33 % of the total variance. The UPGMA phenogram and the PCA (Principle component analysis) indicated that the populations formed five major clusters. CSC 83 (774 g per fruit) and CSC 71 (yellow skin) are considered to be the most important collections to be stressed for further breeding purpose. The available genetic resources of cucumber in Karnataka were characterized.     

Genetic diversity analysis of 60 ginger germplasm core accessions using ISSR and SSR markers

Molecular techniques play a critical role in studies of phylogeny and, thus, have been applied to understand the distribution and extent of genetic variation within and between species. In the present study, a genetic analysis was undertaken using molecular markers (9 ISSR and 13 SSR) on 60 ginger cultivars from different regions of the eastern coast of India (Odisha). The data obtained with 22 polymorphic markers revealed moderate to high diversity in the collection. Both ISSR and SSR markers were efficient in distinguishing all the 60 ginger cultivars. A total of 42 and 160 polymorphic bands were observed with ISSR and SSR markers, respectively. However, SSR markers were observed to be better at displaying average polymorphism (63.29%) than ISSR markers (55%). Analysis of molecular variance results showed that 52 and 66% of the variation occurred among different ginger populations, whereas 48 and 34% of the variation was found within populations, respectively, using ISSR and SSR markers, indicating that ginger cultivars display significant genetic diversity at the population level. Principal coordinates analysis and the dendrogram constructed out of combined data of both markers showed grouping of ginger accessions to their respective area of collection, indicating geographical closeness due to genetic similarity irrespective of the relationship that exists at the morphological level.     

Polyploidy creates higher diversity among Cynodon accessions as assessed by molecular markers

Developing a better understanding of associations among ploidy level, geographic distribution, and genetic diversity of Cynodon accessions could be beneficial to bermudagrass breeding programs, and would enhance our understanding of the evolutionary biology of this warm season grass species. This study was initiated to: (1) determine ploidy analysis of Cynodon accessions collected from Turkey, (2) investigate associations between ploidy level and diversity, (3) determine whether geographic and ploidy distribution are related to nuclear genome variation, and (4) correlate among four nuclear molecular marker systems for Cynodon accessions’ genetic analyses. One hundred and eighty-two Cynodon accessions collected in Turkey from an area south of the Taurus Mountains along the Mediterranean cost and ten known genotypes were genotyped using sequence related amplified polymorphism (SRAP), peroxidase gene polymorphism (POGP), inter-simple sequence repeat (ISSR), and random amplified polymorphic DNA (RAPD). The diploids, triploids, tetraploids, pentaploids, and hexaploids revealed by flow cytometry had a linear present band frequency of 0.36, 0.47, 0.49, 0.52, and 0.54, respectively. Regression analysis explained that quadratic relationship between ploidy level and band frequency was the most explanatory (r = 0.62, P < 0.001). The AMOVA results indicated that 91 and 94% of the total variation resided within ploidy level and provinces, respectively. The UPGMA analysis suggested that commercial bermudagrass cultivars only one-third of the available genetic variation. SRAP, POGP, ISSR, and RAPD markers differed in detecting relationships among the bermudagrass genotypes and rare alleles, suggesting more efficiency of combinatory analysis of molecular marker systems. Elucidating Cynodon accessions’ genetic structure can aid to enhance breeding programs and broaden genetic base of commercial cultivars. O. Gulsen and S. Sever-Mutlu contributed equally to this work.     

Determination of genetic diversity among Saccharina germplasm using ISSR and RAPD markers

Various species of genus Saccharina are economically important brown macroalgae cultivated in China. The genetic background of the conserved Saccharina germplasm was not clear. In this report, DNA-based molecular markers such as inter simple sequence repeats (ISSR) and random amplified polymorphic DNA (RAPD) were used to assess the genetic diversity and phylogenetic relationships among 48 Saccharina germplasms. A total of 50 ISSR and 50 RAPD primers were tested, of which only 33 polymorphic primers (17 ISSR and 16 RAPD) had an amplified clear and reproducible profile, and could be used. Seventeen ISSR primers yielded a total of 262 bands, of which 256 were polymorphic, and 15.06 polymorphic bands per primer were amplified from 48 kelp gametophytes. Sixteen RAPD primers produced 355 bands, of which 352 were polymorphic, and 22 polymorphic bands per primer were observed across 48 individuals. The simple matching coefficient of ISSR, RAPD and pooled ISSR and RAPD dendrograms ranged from 0.568 to 0.885, 0.670 to 0.873, and 0.667 to 0.862, revealing high genetic diversity. Based on the unweighted pair group method with the arithmetic averaging algorithm (UPGMA) cluster analysis and the principal components analysis (PCA) of ISSR data, the 48 gametophytes were divided into three main groups. The Mantel test revealed a similar polymorphism distribution pattern between ISSR and RAPD markers, the correlation coefficient r was 0.62, and the results indicated that both ISSR and RAPD markers were effective to assess the selected gametophytes, while matrix correlation of the ISSR marker system (r = 0.78) was better than that of the RAPD marker system (r = 0.64). Genetic analysis data from this study were helpful in understanding the genetic relationships among the selected 17 kelp varieties (or lines) and provided guidance for molecular-assisted selection for parental gametophytes of hybrid kelp breeding.     

RAPD and ISSR markers in the evaluation of genetic divergence among accessions of elephant grass

Considering the expected genetic variability of elephant grass (Pennisetum purpureum), due to its cultivation in different continents, we characterized and estimated the genetic divergences between 46 accessions of elephant grass with different edaphoclimatic adaptations, using RAPD and ISSR markers. We evaluated, comparatively, the consistency of the information achieved with these markers. Twenty-six RAPD and 25 ISSR primers were employed. The RAPD markers produced 185 bands, 72% of which were polymorphic, with a mean of 5.11 polymorphic bands per primer. The 25 ISSR starters produced 216 bands; 76% were polymorphic, with a mean of 6.56 polymorphic bands per primer. The correlation between the genetic distances achieved by the RAPD and ISSR markers was 0.76, which is highly significant by the Mantel test. Based on UPGMA grouping, considering the point of sudden change, five and six groups were formed for the data from the RAPD and ISSR markers, respectively. These markers provided partially concordant groups, indicating that these techniques can provide consistent information and consequently could be used in studies of genetic diversity among accessions.     

Genetic relationships among Mediterranean Pistacia species evaluated by RAPD and AFLP markers

Polymorphisms among Mediterranean basin Pistacia species and accessions within species were assessed by random amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP) analyses. Twenty-eight Pistacia accessions representing six species from geographically diverse locations in the Mediterranean area were analyzed. With RAPD, a total of 259 DNA fragments were amplified by 27 pre-selected primers, 254 were polymorphic fragments. AFLP analysis with 15 primer sets, produced 954 (93%) polymorphic bands out of a total of 1026. A Mantel test revealed an extremely high correlation (r=0.99) between similarity matrices generated from RAPD and AFLP data sets, indicating that similar results were obtained by the two techniques. Dendrograms constructed from the similarity matrices showed that Pistacia species could be clustered into two groups, one group containing all the #E5/E5#. lentiscus and the second group containing all other accessions. The latter group was divided into two subgroups, one consisting of #E5/E5#. palaestina and #E5/E5#. terebinthus; the other consisting of #E5/E5#. atlantica, #E5/E5#. khinjuk and #E5/E5#. vera. P. vera and P. khinjuk were highly similar, as were P. palaestina and P. terebinthus.     

Genetic diversity in European and Mediterranean faba bean germ plasm revealed by RAPD markers

Broadening of the genetic base and systematic exploitation of heterosis in faba bean requires reliable information on the genetic diversity in the germ plasm. Three groups of faba bean inbred lines were examined by means of RAPDs (random amplified polymorphic DNAs) assays: 13 European small-seeded lines, 6 European large-seeded lines, and 9 Mediterranean lines. Out of 59 primers, 35 were informative and yielded 365 bands, 289 of which were polymorphic with a mean of 8.3 bands per primer. Monomorphic bands were omitted from the analyses and genetic distances (GD) were estimated via the coefficient of Jaccard. The mean GD among the European small-seeded lines was significantly greater than those among the lines of the other two groups. Repeatability of GD estimates was high. Cluster (UPGMA) and principal coordinate analyses identified European small-seeded lines and Mediterranean lines as distinct groups with European large-seeded lines located in between. The results are in harmony with published archaeobotanical findings. We conclude that RAPDs are useful for classification of germ plasm and identification of divergent heterotic groups in faba bean.     

Genetic relationships among Stylosanthes species as revealed by sequence-tagged site markers

 Nineteen sequence-tagged site (STS) primer pairs were designed on coding and non-coding regions in nine published Stylosanthes genes, which were mostly derived from cDNA. Direct sequencing of PCR products derived from genomic DNA allowed us to identify introns and to design specific primers flanking these introns. The use of 24 STS primer pairs for the detection of intra- and inter-specific variation in Stylosanthes based on size differences was tested on a core set of Stylosanthes species. Based on these results, 20 STS markers were selected to determine genetic relationships among 63 genotypes representing 24 Stylosanthes species. A total of 148 alleles were amplified and analyzed, resulting in a genetic similarity value ranging from 0.62 to 0.98 among the species. Based on cluster analysis, three main groups and three subgroups were determined, and most of the species were classified unambiguously. Alloploid species were recognized by the occurrence of more than one allele per STS marker, indicating fixed heterozygosity. Sixteen STS markers were useful for the identification of genotypes within a species. Inter-species relationships, as revealed by STS, were in general agreement with previous morphological and molecular relationship studies. These STS markers are useful as an additional tool for the identification of species, subspecies and genotypes in Stylosanthes, with a view to plant conservation and breeding. Received: 2 June 1998 / Accepted: 28 October 1998     

Genetic diversity and relationships among germplasm of Jatropha curcas L. revealed by RAPDs

Variation is the phenomenon where individuals of a population differ from each other. The variation or diversity can have morphological manifestation or genetic basis. Individual identification based on morphological record is the most common practice in Jatropha. Therefore, in order to find a proper method for estimation of genetic diversity and genetic relationships among different germplasm of Jatropha curcas L., random amplified polymorphic DNA (RAPD) technique based on polymerase chain reaction (PCR) was used for described purpose. Out of 55 decamer primers tested, 26 primers produced good amplification products. A total of 6,011 amplification products were scored from which only 1,859 bands (30.92%) were found to be polymorphic and the size of bands ranged from 300 to 2,500 bp. Unweighted pair group method using arithmetic average cluster analysis revealed clear genetic difference among J. curcas germplasm. The scientific data presented in this study suggests that RAPD-PCR could be used as a valuable tool for estimation of genetic diversity and genetic relationship among germplasm of J. curcas L.     

Genetic diversity in important members of Cucurbitaceae using isozyme, RAPD and ISSR markers

Biochemical and molecular markers have been used on eleven species of Cucurbitaceae collected from lower Gangetic plains. Six enzyme systems were selected. Among 40 primers examined, 14 random amplified polymorphic DNA (RAPD) and 10 inter-simple sequence repeat (ISSR) primers were selected for the analysis. Generated RAPD (100) and ISSR (100) fragments showed high variations among the species. Jaccard similarity coefficients were used for the evaluation of pairwise genetic divergence; cluster analysis of the similarity matrices was performed to estimate interspecific diversity. Further, principal coordinate analysis was performed to evaluate the resolving power of the three marker systems to differenciate among the species.     

Genetic diversity in Bambara groundnut (Vigna subterranea L.) germplasm revealed by RAPD markers.

Random amplified polymorphic DNA (RAPD) markers were used to assess genetic diversity in Bambara groundnut (Vigna subterranea L.) germplasm using 25 African accessions from the collection in the International Institute for Tropical Agriculture, Ibadan, Nigeria. Fifty random decamer primers were screened to assess their ability to detect polymorphism in bambara; 17 of them were selected for this study. Considerable genetic diversity was found among the V. subterranea accessions studied. The relationships among the 25 accessions were studied by cluster analysis. The dendrograms showed two main groups of accessions mainly along the lines of their geographic origin. It is concluded that RAPD can be used for germplasm classification in bambara groundnut and hence for improving this crop.