Taxonomic status of Oryza glumaepatula Steud. II. Hybridization between New World diploids and AA genome species from Asia and Australia

Intraspecific and interspecific crosses were made using O. glumaepatula Steud., O. rufipogon Griff., O. nivara Sharma et Shastry, O. meridionalis Ng, and other diploid accessions from the New World to determine biosystematic relationships among the New World, Asian, and Australian AA genome species. All intraspecific and interspecific combinations produced seeds and hybrids, but at different levels of success. The O. glumaepatula, O. rufipogon, and O. nivara intraspecific hybrids were generally fertile with mean pollen stainability ranging from 75.2% to 80.1% and mean spikelet fertility varying from 32.1% to 87.7%. The O. meridionalis intraspecific crosses showed 3.8% pollen stainability and 0.1% spikelet fertility. Interspecific hybrids showed varying fertilities. Crosses of O. glumaepatula with the New World diploid accessions IRGC 103812 and 105561 produced highly fertile hybrids with 68.0% to 89.7% pollen stainability and 70.8% to 86.6% spikelet fertility. In crosses between O. glumaepatula and the Asian species and other diploid New World accessions IRGC 100961, 103810, and 104386, sterile hybrids were produced with pollen fertility ranging from 0 to 35.1% and spikelet fertility from 0 to 8.2%. IRGC 103810 and 104386 formed fertile hybrids when crossed to O. nivara and O. rufipogon, which were also interfertile. Interspecific crosses of O. meridionalis with the other species produced highly sterile hybrids.     

Taxonomic status of Oryza glumaepatula Steud. I. Comparative morphological studies of New World diploids and Asian AA genome species

This study was carried out to clarify the taxonomic status of the New World diploid wild rice species, Oryza glumaepatula. The morphological variation of 26 diploid rice accessions in the International Rice Genebank at IRRI from South America and Cuba was compared with that of O. rufipogon and O. nivara from Asia. The 28 morphological characters included 16 spikelet and grain, eight leaf and culm, and four panicle characters, and were analyzed using principal component analysis (PCA) and hierarchical agglomerative cluster analysis. The first two principal components accounted for 53.6% of the total variation. The first component was characterized by leaf and sterile lemma characters, 5-panicle dry weight, and grain length, and the second by anther length and its ratio to grain length, grain width, thickness, and 20-hulled grain weight. The characters effectively differentiated the wild rices from Surinam, French Guiana, and the lower Amazon River basin in Brazil, which showed high negative scores along both axes, from other accessions from Venezuela, Colombia, and Cuba, and the Asian species. Most of the accessions from South America are quite distinct from O. rufipogon, with which they have often been grouped as a single species in some taxonomic treatments. Groupings obtained from cluster analysis corresponded closely with the results from PCA. This study supports a distinct taxonomic status of a group of diploid wild rices from South America as O. glumaepatula.     

Hybridization of AA genome rice species from Asia and Australia I. Crosses and development of hybrids

Interspecific and intraspecific crosses of rice species were made involving the AA genome wild species Oryza meridionalis Ng from Australia, O. nivara Sharma et Shastry, O. rufipogon Griff, the weedy type O. sativa f. spontanea Rosch., and the cultigen O. sativa L. from Asia. Seed set and the number of hybrids obtained from both interspecific and intraspecific crosses were low and no significant differences were observed between the interspecific and intraspecific crosses. In the combination O. meridionalis × O. nivara and O. meridionalis × O. rufipogon, considerable differences in reciprocal crosses were observed, whereas higher seed set and more hybrids were obtained when O. meridionalis was used as the female parent. Pollen stainability and seed fertility of hybrids were very low, averaging between 1% and 9% in both interspecific and intraspecific combinations, except in the O. nivara intraspecific cross of 105386 × 106111, whose hybrid showed 83.0% pollen stainabi lity and 66.5% seed fertility.     

Hybridization of AA genome rice species from Asia and Australia II. Meiotic analysis of Oryza meridionalis and its hybrids

To determine the genomic constitution of Oryza meridionalis Ng (2n=2x=24) and to estimate genomic affinity between Asian and Australian wild species of rice containing the AA genomes, chromosome pairing was analyzed at metaphase I in O. meridionalis, O. rufipogon Griff. (2n=2x=24), O. nivara Sharma et Shastry (2n=2x=24), O. sativa f. spontanea Rosch, and their artifical hybrids. The Oryza parental species and their F1 hybrids showed normal meiosis, but slightly reduced chiasma fequency was observed in the hybrids. It is concluded from the cytological analysis that (i) the Australian O. meridionalis contains the AA genome which has very high affinity to that in the Asian AA genome wild species of rice; (ii) chromosome structual changes, such as inversions and reciprocal translocations, have occurred in the genomes of the different species studied.     

Genetic Differentiation in <Emphasis Type="Italic">Oryza meridionalis</Emphasis> Ng based on Molecular and Crossability Analyses

Molecular and hybridization studies were carried out to detect variation patterns in O. meridionalis. A total of 119 polymorphic RAPD markers were produced from 12 10-mer operon primers while 67 alleles were detected using 11 SSR primers. Cluster analysis based on RAPD and SSR markers identified distinct clusters for accessions coming from Irian Jaya (Indonesia) and Queensland, the Northern Territory, and Western Australia. Intraspecific hybrids showed pollen stainability and panicle fertility ranging from 0 to almost 97%. Fertile hybrids were obtained from crosses between accessions of the same geographic origin, specifically those involving the Irian Jaya accessions that showed greater than 70% pollen stainability and panicle fertility. In crosses involving accessions with different origins, partially fertile to sterile hybrids were obtained between accessions coming from Irian Jaya and Queensland. In contrast, most crosses between the Northern Territory and Irian Jaya accessions and Northern Territory and Queensland accessions produced sterile hybrids. The study proves that O. meridionalis is a good taxonomic species undergoing gradual speciation corresponding to its geographic distribution in northern Australia and Irian Jaya.     

Interspecific rice hybrid of Oryza sativa x Oryza nivara reveals a significant increase in seed protein content

Wild species offer a potential reservoir of genetic variation for crop improvement. Besides the valuable genes for disease resistance that the wild species have provided for rice improvement, recent studies have shown that these wild species could also provide favorable alleles for the improvement of yield and yield-related traits. The present study reports yet another potential of wild relatives of rice, which involves the improvement of seed protein content. A significant increase in seed protein content was observed in an interspecific hybrid between Oryza sativa ssp. indica and the wild species Oryza nivara. The hybrid showed a protein content of 12.4%, which was 28 and 18.2% higher than those of the parents O. nivara and IR 64, respectively. The increase in protein content was dependent on the genetic background of the rice variety used in the hybridization. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of seed storage proteins demonstrated that a significant increase in prolamins and glutelins was mainly responsible for the elevated protein content of the hybrid. Amino acid analysis of seed proteins revealed that the hybrid had net gains of 19.5% in lysine and 19.4% in threonine over the O. nivara parent on a seed dry weight basis. Molecular analysis indicated that the increase in protein content of the hybrid was not a result of chromosomal rearrangements or transposable element activation, at least in the chromosomal regions containing seed storage protein genes. A preliminary genetic analysis of the F 2 segregating population showed that the inheritance of the increased protein content was polygenic in nature. The development of this interspecific hybrid offers a great potential for selecting new rice cultivars that combine the high yield and superior cooking quality of IR 64 with improved seed protein content.     

Identification of genomic constitutions of <Emphasis Type="Italic">Oryza</Emphasis> species with the B and C genomes by the PCR-RFLP method

Oryza officinalis complex is the largest and the most complicated group in the genus Oryza L., consisting of about ten species with the B, C, BC, CD, and E genomes. Taxonomy and identification of the species, particularly those with the B, C and BC genomes, are difficult due to the similar morphology and overlapping distribution of some species. The difference in ploidy levels of some species adds more complexity. In the present study, we surveyed 64 accessions of rice germplasm in the O. officinalis complex using RFLP analysis of PCR-amplified Adh genes in addition to chromosome counting. The results confirmed that all O. rhizomatis accessions are diploids with the C genome, whereas all O. minuta accessions are tetraploids having the BC genome. However, both diploid and tetraploid forms were found for the accessions identified in the genebank as O. officinalis, O. punctata and O. eichingeri. The tetraploid form of O. officinalis with the BC genome is exclusively distributed in India and has been described as O. malampuzhaensis. The tetraploid form of O. punctata which has been called O. schweinfurthiana by some workers was found to be as widely distributed as its diploid form in Africa. It is noteworthy that two accessions that had been determined as tetraploid O. officinalis were actually belonging to a species with the CD genome (O. latifolia). Our results promote a better understanding of the genomic constitutions of many accessions in the O. officinalis complex and correct determination of the genebank material, which serves as an important basis of germplasm cataloguing for further research and utilization.     

Electrophoretic variation in seed proteins and interrelationships of species in the genus <Emphasis Type="Italic">Oryza</Emphasis>

Comparative studies of two cultivated and sixteen wild species of the genus Oryza were carried out using one- and two-dimensional gel electrophoresis for variation in their seed proteins for interrelationships of these species. A number of polypeptides in the range of molecular weight 13–110 kDa were seen to vary. Under reducing conditions, polypeptides spread over the regions of mol. wt. 33–40.5, 25–27 and 19–21.5 kDa exhibited maximum variation in their patterns. Two-dimensional gel electrophoresis revealed the occurrence of disulphide-linked glutelin polypeptide pairs of mol. wt. 60, 58, 52 and 25 kDa breaking into a large and a small subunit each in the range of mol. wt. 18–40.5 and 16–25 kDa respectively in Oryza sativa. The number of such polypeptide pairs varied from 2 to 6 in different species and also in O. sativa showed variation in mol. wts. of their constituent subunits. The UPGMA dendrogram revealed that most of the Oryza species occurred in different clusters and subclusters and thus did not share very close relationships. The undisputed and closest relationship observed was that of cultivated rice O. sativa with the O. rufipogon followed by that with O. nivara. The African cultivated O. glaberrima occurring on the nearest branch of the same subcluster, thereby, supporting the phylogenetic of these species suggested in earlier studies. Eight diploid species and seven tetraploid species were included in one part of the dendrogram while the remaining two species with AA genome i.e. O. glumaepatula and O. meridionalis and one with FF i.e. O. brachyantha stood separately from these as scattered in the group of seven tetraploid species with BBCC, CCDD and HHJJ genomes. The tetraploids O. alta, O. latifolia and O. grandiglumis with CCDD genomes which occurred on the farthest part were distantly related with O. sativa. The cyanogen bromide peptide maps and two dimensional gel electrophoresis also supported the closest relationship between O. sativa and O. rufipogon.     

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 Accessions of Wild Rice Oryza granulata from South and Southeast Asia

Oryza granulata, an upland wild rice species, represents an unique germplasm for possessing abilities of tolerance to shade and drought, immune to bacterial blight and resistance to brown planthopper. Although low degree of genetic variability has been revealed within its populations, little genetic information at the species level is available in determining rational conservation strategies. Here we used dominant DNA marker random amplified polymorphism DNA (RAPD) to assess the genetic variability among 23 accessions of O. granulata that collected from main distribution areas worldwide. Twenty decamer primers generated a total of 243 bands, with 83.5% of them (203 bands) being polymorphic. Calculation of Shannon index of diversity revealed an average value of 0.42 ± 0.25, indicating that O. granulata maintains a relatively high degree of genetic diversity on the species level. Analysis of genetic dissimilarity (GD) showed that genetic differentiation occurred among studied accessions, which supports the feasibility of current ex situ conservation strategies. We also suggested that information based on population studies, which could be achieved by international co-operation, is needed to conserve this widespread germplasm more effectively.     

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.     

Geographic Pattern of Genetic Diversity Among 43 Ethiopian Mustard(Brassica carinata A. Braun) Accessions as Revealed by RAPD Analysis

As an oilseed crop, the cultivation of Ethiopian mustard (Brassica carinata) is restricted only to Ethiopia. Even though geographic diversity is a potent source of allelic diversity, the extent of genetic diversity among germplasm material of Ethiopian mustard from different countries has not been assessed. Forty-three accessions, comprising 29 accessions from eight different geographic regions of Ethiopia and 14 exotic accessions from Australia, Pakistan, Spain, and Zambia were analysed for their genetic diversity using random amplified polymorphic DNA (RAPD) technique. A set of 50 primers yielded a total of 275 polymorphic bands allowing an unequivocal separation of every Ethiopian mustard accession. The usefulness of the 50 RAPD primers in measuring heterozygousity and distinguishing accessions was variable such that polymorphic information content (PIC) varied from 0.05 to 0.40, band informativeness (BI) from 0.05 to 0.65 and primer resolving power (RP) from 0.15 to 6.83. Jaccard's similarity coefficients ranged from 0.44 to 0.87 indicating the presence of a high level of genetic diversity. On the average, Australian and Ethiopian accessions were the most similar while, Spanish and Zambian accessions were the most distant ones. Cluster analysis grouped the 43 accessions into four groups, which has quite a high fit (r = 0.80) to the original similarity matrix. With no prior molecular information, the RAPD technique detected large genetic diversity among the 43 accessions from five different countries and their grouping by dendrogram and principal coordinate analysis (PCoA) was inclined towards geographic differentiation of RAPD markers. Conversely, RAPD differentiation along geographic origin was not apparent within the Ethiopian accessions.     

Genetic variation in common and runner bean of the Northern Meseta in Spain

The genetic variation within and between Spanish landraces or varieties of Phaseolus vulgaris L. (common bean) and P. coccineus L. (runner bean) has been estimated by means of isozymes and random amplified polymorphic DNA (RAPD) analyses. Likewise, storage protein and amino acid content in dry seeds have been estimated. Fifteen landraces (60 accessions) of P. vulgaris and six of P. coccineus (six accessions) have been studied. Of the seven isozymatic systems analyzed only three systems and three loci showed variability in each species. Isozyme analyses revealed that genetic variability within and between landraces exist in both species. Even variability within accession was detected in some P. vulgaris landraces. Comparison of isozyme data indicated that Spanish landraces have a lower level of genetic variability than wild American materials and probably also lower than American landraces. RAPD analysis allowed for the uniquely distinguishing of all landraces. Genetic similarity among landraces, estimated by both isozymes and RAPDs, were not related with the seed morphological characters (color, size and shape) which define each variety or landrace. Variation in protein and amino acid content among landraces was also detected. The average protein content in common bean (20.48%) was similar to values previously reported in this species and higher than the average in the runner bean landraces (16.33%). In relation to the amino acid content methionine and cysteine showed the lowest values in all samples, although the content of these two amino acids varied widely among landraces.     

Genetic variability evaluation in a Moroccan collection of barley, Hordeum vulgare L., by means of storage proteins and RAPDs

The genetic variation existing in a set of barley (Hordeum vulgare L.) landrace samples recently collected in Morocco was estimated. Two kinds of genetic markers, seed storage proteins (hordeins) and random amplified polymorphic DNA (RAPD), were used. Only six out of 31 landraces were subjected to RAPD analysis. Both kinds of markers, RAPD and storage proteins, yielded similar results, showing that the level of variation observed in Moroccan barley was high: all landraces showed variability; 808 different storage protein patterns (multilocus associations) were observed among 1897 individuals (2.32 seeds per association, on average) with an average of 43 multilocus associations per accession. In general, genetic variation within accessions was higher than between accessions. The 100 polymorphic RAPD bands generated by 21 effective primers were able to generate enough patterns to differentiate between uniform cultivars and even between individuals in variable accessions. One of the aims of this work was to compare the effectiveness of RAPD versus storage protein techniques in assessing the variability of genetic resource collections. On average hordeins were more polymorphic than RAPDs: they showed more alternatives per band on gels and a higher percentage of polymorphic bands, although RAPDs supply a higher number of bands. Although RAPD is an easy and standard technique, storage protein analysis is technically easier, cheaper and needs less sophisticated equipment. Thus, when resources are a limiting factor and considering the cost of consumables and work time, seed storage proteins must be the technique of choice for a first estimation of genetic variation in plant genetic resource collections.     

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 in cowpea [Vigna unguiculata (L.) Walp.] as revealed by RAPD markers

The present study, using RAPD analysis, was undertaken to characterize genetic variation in domesticated cowpea and its wild progenitor, as well as their relationships. The materials used consisted of 26 domesticated accessions, including accessions from each of the five cultivar-group, and 30 wild/weedy accessions, including accessions from West, East and southern Africa. A total of 28 primers generated 202 RAPD bands. One hundred and eight bands were polymorphic among the domesticated compared to 181 among wild/weedy cowpea accessions. Wild accessions were more diverse in East Africa, which is the likely area of origin of V. unguiculata var. spontanea. Var. spontanea is supposed to have spread westward and southward, with a loss of variability, loss counterbalanceed in southern Africa by introgressions with local perennial subspecies. Although the variabilty of domesticated cowpea was the highest ever recorded, cultivar-groups were poorly resolved, and several results obtained with isozyme data were not confirmed here. However primitive cultivars were more diverse than evolved cultivars, which still suggests two consecutive bottlenecks within domesticated cowpea evolution. As isozymes and AFLP markers, although with a larger number of markers, RAPD data confirmed the single domestication hypothesis, the gap between wild and domesticated cowpea, and the widespread introgression phenomena between wild and domesticated cowpea.     

Evaluation of Antioxidant,Lipid, and Protein Fractions of Accessions of Oryza Species

The genus Oryza has given rise to rice (Oryza sativa L.), a major source of food for much of the human population. The Oryza genus is small, including only 23 species, but it is remarkably diverse in terms of its ecological adaptation. This diversity may not be only restricted to ecological characteristics but also to kernel end‐use quality characteristics. This study was undertaken to evaluate Oryza species as a gene pool for improving the properties of rice bran for human consumption. Several accessions of 13 Oryza species were grown in a greenhouse, along with eight rice (Oryza sativa L.) accessions displaying low and high bran oil content, as well as low and high palmitic acid content. The total lipid content of the Oryza species was within the range found for Oryza sativa accessions. However, the level of palmitic acid in the O. species was as high as that reported for soybean high‐palmitic acid mutants. Oryza species also contained higher levels of the γ‐oryzanol and phenolic fractions compared to cultivated rice. Low or not significant phenotypic correlations between lipid, palmitic acid, and γ‐oryzanol suggest that these fractions in cultivated rice could be increased simultaneously using several of the Oryza species accessions identified in this study. A cultivar with enhanced levels of these fractions would be suitable for use in the production of high γ‐oryzanol margarine, shortening, and frying oils.     

Marker-assisted sampling of the cultivated Andean potato Solanum phureja collection using RAPD markers

The potato crop originated in the Andean highlands where numerous farmer's varieties and non-cultivated wild species exist. An Andean potato collection is held in trust at the International Potato Center (CIP) to preserve the biodiversity of this crop and ensure the supply of germplasm for potato improvement worldwide. A core collection representing the biodiversity of the Andean potato germplasm is under construction using morphological, molecular, and geographic data. One of the eight cultivated potato species, Solanum phureja, has been genotyped using the RAPD technique. A protocol suitable for large germplasm collection genotyping has been developed to process numerous samples at reasonable costs. From 106 RAPD primers evaluated, we have selected 12 primers yielding 102 polymorphic markers, which unambiguously discriminated all 128 accessions but 2 that are possible duplicates. The S. phureja germplasm collected throughout the Andean countries appears to have a homogeneous genetic constitution. There was no clear geographic pattern as indicated by cluster analysis of the RAPD data. A sub-group of 20 accessions has been identified on the basis of the marker data and selected to maximize molecular (RAPD) variance and polymorphism. The probability of capturing equal amounts of marker polymorphism in this sub-group of 20 accessions by random sampling is less than 40%. This set accessions represents our first group of accessions that may constitute a core of the S. phureja collection. This tentative core will be challenged for diversity content by alternate markers and agronomic traits. Hence, the methodology for sampling less than 10% of the base collection, proposed for core collections by Brown (1989), can be based on molecular marker data provided cost-efficient fingerprints are developed.     

Assessment of genetic diversity and genetic relationships among 29 populations of <Emphasis Type="Italic">Azadirachta indica</Emphasis> A. Juss. using RAPD markers

Randomly amplified polymorphic DNA (RAPD) analysis was employed to assess genetic divergence among 29 neem accessions collected from two agro-ecological regions of India (11 agro-climatic sub-zones), which cover three states, Punjab, Haryana and Rajasthan. Out of 24, 10-mer random primers used for studying genetic divergence, 14 were polymorphic, generating a total of 73 amplification products with an average of 5.21 products per polymorphic primer and estimated gene diversity of 0.49. Genetic relationships among accessions were evaluated by generating a similarity matrix based on Jaccard’s coefficient, ranging from 0.70 to 0.96. The phenetic dendrogram generated by UPGMA analysis grouped accessions into five clusters. RAPD performed within accessions (individual seedlings collected from the same mother plant) showed no variation indicating homogeneous population within accessions. Primers OPA-18, OPC-08 and OPI-03 were found most informative based on their resolving power. The degree of genetic variation detected among the 29 accessions with RAPD analysis suggests that RAPD can be used for studying genetic diversity in neem. The study also demonstrated that neem germplasm collected from northwestern plains of India shows no eco-geographical isolation based on sub-zones because accessions collected from different sub-regions are grouping together in the genetic tree.