Estimating the genetic relationship of hybrid bromegrass to smooth bromegrass and meadow bromegrass using RAPD markers

The two most widely grown bromegrass species in North America are smooth bromegrass, a hay type grass, and meadow bromegrass, a pasture type grass. Hybrid populations between these two species have been developed through hybridization and recurrent selection. The objective of this study was to determine the genetic relatedness of the hybrid bromegrass population S‐9073M to its parental populations using random amplified polymorphic DNA (RAPD) markers and to determine genetic variation within and between populations. Individuals from each of the three populations were genotyped at 43 polymorphic RAPD loci. One of the RAPD fragments was meadow bromegrass‐specific. Cluster analysis showed three groups representing the two parental populations and the hybrid population. An analysis of molecular variance (AMOVA), showed that the hybrid population had the highest within‐population variation, followed by meadow bromegrass and smooth bromegrass. The interpopulation genetic distance (phi‐statistic =Φ_st) was highest between meadow and smooth bromegrass and lowest between smooth and hybrid bromegrass. The hybrid population was genetically intermediate to smooth bromegrass and meadow bromegrass, but closer to smooth bromegrass, possibly reflecting the selection criteria used in the development of this hybrid.     

Genetic variation, heritability and progeny testing in meadow bromegrass

Meadow bromegrass is a recently introduced pasture grass in western Canada. Its leafy production and rapid regrowth have made it a major grass species for pasturing beef animals in this region. Because relatively little breeding work has been done on this species, there is little information on its breeding behaviour. The main objective of this study was to estimate genetic parameters and assess breeding methodologies for meadow bromegrass. Forty half‐sib (polycross and open‐pollinated) and full‐sib (selfed) progenies were evaluated for dry matter and seed yield, fertility index, harvest index, plant height, plant spread, crude protein, neutral detergent fibre and acid detergent fibre. Genetic variation for seed yield, harvest index, and plant height was significant in the open‐pollinated and selfed tests. Genetic variation estimates for dry matter yield were negative (polycross), not significant (open‐pollinated), or significant (selfed). Estimates of genetic variation for quality traits were not significant, except for acid detergent fibre (selfed). Correlations among characters indicated that it is possible to simultaneously improve seed and forage yield. Rankings of progenies by the half‐sib tests for forage and seed yield were not altered by the use of the synthetic parental value (SV_i), which includes information from selfed progeny. The polycross progeny test did not discriminate parents as well as the open‐pollinated and selfed tests; this may have been a result of non‐random pollination in the polycross. Correlation among the progeny tests, showed that open‐pollinated and selfed progeny tests agreed for all characters, except for harvest index, while correlation between polycross and selfed tests were significant for six characters but not dry matter yield, and acid and neutral detergent fibres. The most dissimilar tests were polycross and open‐pollinated, with significant correlations found only for fertility index, height, spread and neutral detergent fibre. It was concluded that there was significant variation and moderate heritability for most traits in the meadow bromegrass populations evaluated, and that the open‐pollinated progeny test is the method of choice for selecting parents for synthetics of this species.     

Random amplified polymorphic DNA variation within and among bean landrace mixtures (Phaseolus vulgaris L.) from Tanzania

Genetic characterization of 51 individual pure lines from 13 landraces of three common bean (Phaseolus vulgaris L.) mixtures from the southern highlands of Tanzania was undertaken using random amplified polymorphic DNA (RAPD) analysis. A dendrogram generated by cluster analysis from data derived from fragments amplified by 12 random 10-base primers divided the bean individuals onto two main branches with less than 60% genetic similarity. Branches A and B subdivided into two and four clusters, respectively. Mixture 2, comprising three landraces, was the most uniform, most plants appearing on cluster 4 of branch B. Three of the four landraces of mixture 1 appeared on cluster 3 of branch B while the fourth landrace appeared on major branch A. Mixture 3 showed the greatest genetic variation with components appearing on both major branches. The clear separation of the 13 landraces onto two main branches of the dendrogram together with phenotypic characters, notably variation in bean size, suggests that the two groups might represent two distinct gene pools of P. vulgaris. This revised version was published online in July 2006 with corrections to the Cover Date.     

Random amplified polymorphic DNA variation among German and Dutch asparagus cultivars

DNA polymorphism among nine cultivars of Asparagus officinalis L. was measured using random amplified polymorphic DNA (RAPD). Of 69 reproducible amplification products from 12 arbitrary decamer primers, 49 RAPD markers were polymorphic and could be used to distinguish six German and three Dutch asparagus cultivars. Even with very small sample sizes, genetic similarity measurements based on the RAPD data allowed accurate grouping of the nine cultivars into distinct clusters, with the exception of two individuals which clustered to closely related varieties. Two German cultivars showed high genetic similarity and were distinct from the remaining German varieties. The German and Dutch cultivars were clearly separated by a relatively large genetic distance.     

Evaluating genetic variation and relationships among two bromegrass species and their hybrid using RAPD and AFLP markers

The two most widely grown bromegrass species in North America are smooth bromegrass (Bromus inermis Leyss.), a hay type grass, and meadow bromegrass (Bromus riparius Rehmann), a pasture type grass. Recently a hybrid bromegrass population between the two species has been produced as a dual-purpose hay-pasture grass. Molecular markers have the potential to improve selection procedures to enhance bromegrass breeding. The objective of this study was to use RAPD and AFLP markers to determine genetic relationships and variations among bromegrass populations. Forty-three RAPD markers from 21 primers and 83 AFLP markers from seven primer combinations were used. Both marker types were able to group the individuals into their respective populations. The relationships among the individuals within each of the populations were not similar between the two marker types. Analysis of molecular variance (AMOVA) detected greater within-population variation than among-population variation for both marker types. The highest variation was observed in the hybrid population followed by meadow and then smooth bromegrass. The inter-population distance from both markers indicated that the highest genetic distance was between meadow and smooth bromegrass and lowest between smooth and hybrid bromegrass, which reflect the breeding history of the hybrid population. This study showed that both markers are capable of differentiating bromegrass genotypes into their respective populations, detecting genetic variation and relationships of the populations. Results of this study suggest that these two markers can be used in the future to enhance the current breeding practices in bromegrass, however, AFLP markers would be the marker of choice due to the high number of polymorphic markers. This revised version was published online in July 2006 with corrections to the Cover Date.     

Genetic variation and cultivar identification of Jamaican yam germplasm by random amplified polymorphic DNA analysis

Summary We have used random amplified polymorphic DNA (RAPD) analysis to characterize eleven cultivars of the five economically most important yam species grown in Jamaica (Dioscorea alata, D. cayenensis, D. rotundata, D. trifida and D. esculenta). Amplification of genomic DNA samples with nine different arbitrary 10mer primers revealed a total of 338 different band positions, ranging in size from 0.3 to 2.5 kb. RAPD patterns proved to be highly reproducible and somatically stable. While no variation was observed among plants belonging to the same cultivar, a large number of intervarietal and interspecific polymorphisms enabled us to reliably discriminate between all Jamaican cultivars investigated.     

Assessment of natural and induced genetic variation in Alstroemeria using random amplified polymorphic DNA (RAPD) markers

Summary We have used random amplified polymorphic DNA (RAPD) markers to study genetic variation in Alstroemeria. The first objective was to examine the discriminatory power of RAPD markers in different genotypes of Alstroemeria obtained by traditional breeding. All genotypes examined, including commercial Alstroemeria varieties, could be distinguished on the basis of their RAPD profiles. Progeny plants could be distinguished from their parents. A second objective of this study was to investigate whether RAPD markers can be used as a routine tool to detect mutant plants, as an alternative to glasshouse testing. To address this objective, we analysed Alstroemeria plants that carried phenotypically visible mutations that either were induced by irradiation using X-rays or were the result of somaclonal variation. In eight out of a total of 13 mutant Alstroemeria plants obtained after irradiation or tissue culture we detected no polymorphisms when compared to control plants that were considered to be non-mutated. Only in five of the mutant plants analysed we detected one to two polymorphisms. These results suggest that frequent genome rearrangements had not occurred in the mutant plants analysed. These results also demonstrate that the RAPD technique is an inappropriate tool for the rapid screening of Alstroemeria for induced variation. It that the RAPD technique is an inappropriate tool for the rapid screening of Alstroemeria for induced variation. It seems probable that this conclusion would be equally applicable in other plant genera in which induced variation has occurred. However, the RAPD technique is a simple and effective tool for genetic fingerprinting of Alstroemeria varieties, provided their differences are due to sexual propagation.     

Random amplified polymorphic DNA (RAPD) variation in Fagopyrum tataricum Gaertn. accessions from China and the Himalayan region

The diversity among 52 landraces and cultivars of tartary buckwheat (Fagopyrum tataricum Gaertn.) and one accession of its wild ancestor, F. tataricum ssp. potanini Batalin, from diverse geographic origins was examined using random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) markers. Eighteen primers produced a total of 240 fragments, of which 153 (63.75%) were monomorphic and 87 (36.25%) polymorphic bands. UPGMA-based pairwise Jaccard’s coefficient of similarity was used to deduce the relationships among 53 genetically diverse accessions. The similarity between cultivated tartary buckwheat accessions ranged from 0.61 to 1.00. Four distinct clusters were formed which corresponded well with the geographic distribution of the tartary buckwheat. Nepalese accessions showed maximum diversity followed by Chinese accessions. Tartary buckwheat accessions from the Himalayan region of northwestern India revealed a narrow gene pool. The wild buckwheat accession did not group with any of the three cultivated tartary buckwheat groups, and formed its own single-entry group. Genetic similarity (0.59) of Chinese buckwheat accessions with the wild ancestor reaffirmed that cultivated tartary buckwheat originated in the Yunnan province of northwestern China. Consistent with some earlier reports, our study demonstrated the usefulness of the RAPD technique for the characterization of plant genetic resources and assessment of diversity between species. This revised version was published online in August 2006 with corrections to the Cover Date.     

Molecular genetic diversity within and among German ecotypes in comparison to European perennial ryegrass cultivars

Perennial ryegrass (Lolium perenne L.) is the most important grass species for temperate grassland agriculture. The level and distribution of genetic variation in gene bank ecotype collections is still largely unknown but of great interest for the planning of breeding programmes. The objectives of this study were to (i) assess the molecular variance and population structure of German ecotypes at the regional and population level, (ii) assign ecotypes to germplasm pools and (iii) compare the relationship between German ecotypes and previously‐investigated European cultivars of perennial ryegrass, A total of 22 ecotypes originating from three geographic areas in Germany. each with a sample size of 20 individual plants, were investigated with 156 polymorphic RAPD markers. Genetic distance among ecotypes ranged from 0,27 to 0.48, An analysis of molecular variance (AMOVA) revealed a much larger variation within populations (71%) than among them (29%). Ecotypes from North Germany were significantly different from those of South and Middle Germany. Thus, two distinct germplasm pools could be identified. The 22 ecotypes and 22 previously investigated cultivars shared 98% of the molecular variance.     

Use of RAPD and microsatellite (SSR) variation to assess genetic relationships among populations of tetraploid alfalfa, Medicago sativa

The level of random amplified polymorphic DNA (RAPD) and microsatellite variation present in four ecotypes and two varieties of alfalfa (lucerne) from Italian and Egyptian germplasm sources was evaluated. A sample of 100 plants from 10 populations was analysed by means of 41 RAPD markers and 37 simple sequence repeat (SSR) markers. Both molecular approaches revealed a high degree of genetic diversity within each of the cultivated populations and enabled each of the plants considered to be uniquely fingerprinted. The genetic relationships among plants and populations were analysed by computing AMOVA (analysis of molecular variance) and F_ST analyses. RAPDs were able to separate the Italian populations from the Egyptian variety. SSRs allowed strong separation of the four Italian alfalfa ecotypes. It was concluded that RAPD and microsatellites could be useful and powerful tools for assessing genetic variation and genetic relationships in tetraploid alfalfa.     

Random amplified polymorphic DNA (RAPD) markers variability among cultivars and landraces of common beans (Phaseolus vulgaris L.) of south-Brazil

To evaluate the variability among cultivars and landraces of common bean(Phaseolus vulgaris L.), 15 cultivars and 18 landraces of common bean (Phaseolus vulgarisL.), a undefined species of Phaseolus,two landraces of Vigna angularis L., and a landrace of soybean (Glycine maxL.), were screened with fifteen oligonucleotide primers in PCR reactions. An average of 20.3 RAPD bands were scored per primer. A total of 304 amplification products were scored of which 88.8% were polymorphic among Phaseolus genotypes. Based on the RAPD markers, four major clusters were formed. Three clusters corresponded to the soybean, to the two Vigna angularis landraces, and to the Phaseolus sp. landrace, respectively. The fourth cluster include all the landraces and cultivars of Phaseolus vulgaris. This large group could be separated into three subgroups that were correlated with the phaseolin patterns and the average seed weight of the genotypes. The analysis shows that most of the landraces collected in South Brazil (17 out of 18) belong to the Andean gene pool, and most of the cultivars (13 out of 15) belong to the Middle American gene pool. This revised version was published online in July 2006 with corrections to the Cover Date.     

Random amplified polymorphic DNA analysis of Australian rice (Oryza sativa L.) varieties

Summary The genetic relationships between rice varieties were analysed by using the polymerase chain reaction (PCR), with arbitrary oligonucleotide primers in the random amplified polymorphic DNA (RAPD) method. PCR with 22 arbitrary primers applied to 37 varieties produced 144 useful markers, of which 67% were polymorphic. Thus, with selected primers sufficient polymorphism could be detected to allow identification of individual varieties. Visual examination of electrophoresis gels and analysis of banding patterns confirmed that commercial Australian and USA lines and their relatives were very closely related, with similarity indices of 88–97%. Three varieties originating from more distant geographical centres were easily distinguished, producing variety-specific amplification profiles and expressing a lower similarity index of 80% to all other varieties tested. PCR offers a potentially simple, rapid and reliable method for rice genotype identification and recognition of lines that could contribute genetic diversity to new commercial varieties.Abbreviations PCR Polymerase Chain Reaction - RAPD Random Amplified Polymorphic DNA     

Evaluation of random amplified polymorphic DNA (RAPD) markers in Hevea brasiliensis

The applicability of random amplified polymorphic DNA (RAPD) markers in the cultivated rubber tree, Hevea, was evaluated using 43 decamer oligonucleotide primers in a set of 24 clones selected in different South-East Asian countries. A total of 220 0.35–3.5 kb DNA fragments were amplified, of which 111 were polymorphic. Of these, 80 fragments (RAPD markers) which were repeatable and clearly scorable across all genotypes were used to estimate genetic distances among the clones tested. The estimated genetic distances ranged from 0.05 (RRII 308 and PB 5/51) to 0.75 (RRIC 100 and SCATC 88–13). A mean genetic distance of 0.5 indicates a rather high genetic variability among the tested clones. As expected, because of the breeding history of Hevea, UPGMA cluster analysis and Principal Coordinate Analysis (PCoA) indicated the absence of a distinct geographical grouping. The possible application of RAPD markers for clone identification and also for analysis of genetic relationships among Hevea clones is discussed.     

Polymorphism and DNA markers for asparagus cultivars identified by random amplified polymorphic DNA

Summary DNA polymorphism among five Asparagus officinalis L. cultivars-Imperial, Snow, Steline, UC-157 and Larac, as detected by random amplified polymorphic DNA (RAPD), is reported. Thirty one decamer primers were tested. and twenty six of them yielded amplification products. Fourteen primers gave products with at least one polymorphic DNA fragment. Among a total of 119 amplified fragments 33 were polymorphic. These RAPD markers enabled the identification of asparagus cultivars. Unique markers for cultivars were: Snow-bands 475 bp, 772 bp, 412 bp, 935 bp and 820 bp amplified by primers D5, OPA-07, OPA-09, OPA-10 and OPA-18, respectively. Steline-bands 645 bp, 680 bp and 997 bp amplified by primers A32, OPA-03 and OPA-09, respectively. A band 903 bp, amplitied by primer OPA-12, is a marker for Imperial, and a band 420 bp, amplified by primer D52, is a marker for Larac. Cultivar UC-157 could be identified by a combination of shared polymorphic bands. The pairwise marker difference between cultivars ranged from 0.08 to 0.17. A phenogram of the genetic relationship based on RAPD fits with the known origin of the cultivars.     

Random amplified polymorphic DNA (RAPDs) markers for genetic analysis in micropropagated plants of Robinia pseudoacacia L.

Four years' old micropropagated plants regenerated by enhanced axillary branching from shoot buds of a single genotype of Robinia pseudoacacia were characterized by RAPDs. Random amplified polymorphic DNA analysis was carried outusing 19 random 10-mer DNA primers and 286 RAPD bands were examined which showed 30% polymorphism. Similarity indices ranged from 0.86 to 0.96 among different plants based on RAPD data. The UPGMA dendrogram was constructed based on similarity indices which showed clustering of different plants into subgroups based on similarity values. Our results suggest that somaclonal DNA sequence variations are present even when organized cultures such as shoot buds were used as explant for micro-propagation. This revised version was published online in August 2006 with corrections to the Cover Date.     

Relationships in pineapple by random amplified polymorphic DNA (RAPD) analysis

Random amplified polymorphic DNA (RAPD) analysis was applied to eight commercial cultivars of pineapple, two intergroup hybrids and two wild species. Morphologically, pineapple is divided into the Cayenne, Queen, Spanish, Maipure and Abacaxi groups. Members of the first three groups have been analysed in this study. The cultivars ‘Tradsithong’, ‘Phuket’, ‘Sawee’ and ‘Tainan’, with spiny leaves, form the Queen group. In ‘Pattavia’, ‘Nanglae’ and ‘Petburi no. 2’ (Cayenne group), spines are confined to the leaf tips. ‘Intrachitdang’ is normally placed in the Spanish group, which is morphologically similar to the Queen group, but with inferior quality fruit. DNA amplification products were compared from 16 arbitrary 10‐mer primers from which a dendrogram was constructed. The results confirmed morphological classifications for seven of the eight commercial cultivars, with the Queen and Cayenne groups as separate clusters. However, the cv. ‘Intrachitdang’ was more closely related to the Cayenne group. Two hybrids from reciprocal Cayenne × Queen group crosses, were more closely allied to the Queen group. The two wild species were outside the groups. RAPD analysis can be exploited to investigate relationships within pineapple germplasm.     

Analysis of genetic variation in Cucurbita moschata by random amplified polymorphic DNA (RAPD) markers

Knowledge of genetic relationships among genotypes is essential for the effective utilisation of germplasm, especially for poorly characterised species. Random amplified polymorphic DNA (RAPD) analysis provides a quick and reliable method for resolving genetic relationships. Although Cucurbita moschata Duch, also known as tropical pumpkin, is one of the most important vegetable crops in Africa, being adapted to a wide range of climatic and soil conditions, it is a scientifically neglected species. The objectives of this study were to (1) analyse the amount of genetic diversity inC. moschata landraces grown in south-central Africa and (2) classify the landraces to assist in selection of parent genotypes for improvement of fruit characteristics. Cluster analysis, based on 39 polymorphic and 105 monomorphic DNA fragments amplified by 16 primers, was used to show relationships among 31 genotypes obtained from Zambia and Malawi. The analysis revealed four clusters, with genotypes from Malawi mainly clustering in three clusters while all genotypes from Zambia and three from Malawi clustered in one cluster. The pair-wise mean genetic distance was 0.32 ± 0.04 for samples from Malawi and 0.26 ± 0.04 for samples from Zambia. The possible application of the resulting classification in breeding of C. moschata is discussed. This revised version was published online in August 2006 with corrections to the Cover Date.     

Genetic diversity and identification of cymbidium cultivars as measured by random amplified polymorphic DNA (RAPD) markers

DNA from thirty-six cymbidium cultivars was examined using polymerase chain reaction (PCR) to determine the efficiency of randomly amplified polymorphic DNA (RAPD) markers in identifying cultivars and determining levels of genetic variability. A total of 132 RAPD markers, 78% of which were polymorphic, were produced from 15 10mer arbitrary primers. All the cultivars were distinguishable when a number of primers was considered. One cultivar, Blue Smoke ‘Green Meadow’ could be distinguished from all the rest based only on lack of the OPA5-370 fragment. Genetic distances among the cultivars were estimated based on the amount of band sharing and ranged from 0.08–0.50 with an average of 0.29. Cluster analysis of genetic distance estimates grouped siblings together with each other and parents with offsprings, thereby agreeing with known parentage information and corroborating isozyme data obtained from a separate study. The possible application of the observed polymorphism and variation to cymbidium breeding is discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Random amplified polymorphic DNA (RAPD) analysis for the verification of hybridity in interspecific crosses of Alstroemeria

Random amplified polymorphic DNA (RAPD) markers were used to verify interspecific hybridization in Alstroemeria. Five putative interspecific hybrids and their parents were analysed by means of four preselected RAPD primers. The putative parentage was confirmed in four hybrids and was excluded in one that showed completely different RAPD patterns from its putative parents and a different phenotype. Our results demonstrated that this molecular technique is a powerful tool for verifying hybridity rapidly if the putative parents are given. This tool will allow screening of small immature seedlings for verification of hybridity and should improve the efficiency of breeding programmes.     

RAPD variation within and among populations of globe artichoke cultivar 'Spinoso sardo'

Globe artichoke is a vegetable native to the Mediterranean basin. Its commercial production is mainly based on perennial cultivation of vegetatively propagated clones, which are highly heterozygous and segregate widely when progeny‐tested. The aim of this study was to measure genetic variation using random amplified polymorphic DNA (RAPD) in five populations of ‘Spinoso sardo’, the most widely grown and economically important cultivar in Sardinia (Italy). Analysis of molecular variance (amova ) gave highly significant differences between populations (28.1% of the total genetic diversity); substantial within‐population variation was detected (71.8% of total genetic diversity). The RAPD results show the need to apply clonal selection to narrow the broad genetic variability within the cultivar and to address important questions on the most suitable strategies for germplasm preservation.