A tourist element in the 5'-flanking region of the catalase gene CatA reveals evolutionary relationships among Oryza species with various genome types

Tourist-OsaCatA, a transposable element, was found in the 5′-flanking region of the rice gene CatA. The characteristics of this element are similar to those of the other Tourist elements so far found in Oryza sativa. PCR and sequence analyses of 37 accessions of 18 species revealed that all the Oryza species examined, except for one accession, have either a full-length or a partial Tourist element at this locus. Unlike the Tourist elements previously reported, this Tourist element is found in all four Oryza species complexes in the Oryzeae tribe. All AA genome Oryza species, except O. longistaminata, contain the full-length Tourist element. O. longistaminata and the species of the O. officinalis, O. meyeriana and O. ridleyi complexes contain the partial element. A phylogenetic tree of Oryza species based on the nucleotide sequences of these Tourist elements was constructed. The O. longistaminata accessions were placed near the neighboring cluster of the officinalis complex. We propose that the ancestor of O. longistaminata and that of other species with the AA genome diverged, and the ancestor(s) of the O. officinalis, O. ridleyi and O. meyeriana complexes then diverged from the ancestor of O. longistaminata in the course of the evolution of the Oryza species. The Tourist elements associated with CatA and its orthologs thus provide useful tools for examining evolutionary relationships among Oryza species. Received: 12 March 1999 / Accepted: 7 July 1999     

Phylogenetic relationships among Oryza species revealed by AFLP markers

 The genus Oryza to which cultivated rice belongs has 22 wild species. Seventy-seven accessions of 23 Oryza species, five related genera, and three outgroup taxa were fingerprinted using amplified fragment length polymorphism (AFLP). A total of 1191 polymorphic markers were obtained using five AFLP primer combinations. AFLP data were analyzed to study species relationships using different clustering algorithms, and the resulting phenograms were tested for stability and robustness. The findings suggest a common ancestry to the genus Oryza. Moreover, the results demonstrate that: (1) evolution in Oryza has followed a polyphyletic path wherein multiple lineages underwent independent divergence after separation early in the evolution from a common ancestor/pool of related taxa; (2) newly assigned genomes, GG for O. meyeriana and HHJJ for O. ridleyi complexes, are among the most diverged in the genus; (3) CCDD tetraploids have a relatively ancient origin among the Officinalis complex; (4) O. malampuzhaensis, O. indandamanica, O. alta, and O. grandiglumis are diverged enough to deserve species status; (5) O. officinalis and O. eichingeri (CC) are putative progenitors of O. minuta ^* O. malampuzhaensis and tetraploid O. punctata, respectively, (6) O. brachyantha is most diverged species in the genus. AFLP is reliable molecular technique and provides one of the most informative approaches to ascertain genetic relationships in Oryza, which may also be true for other related species/organisms. Received: 1 July 1998 / Accepted: 2 November 1998     

Structural differences in the vicinity of the waxy locus among the Oryza species with the AA-genome: identification of variable regions

We constructed a fine physical map for a 260-kb rice BAC contig surrounding the waxy locus. In order to identify variable regions within this 260-kb as to the restriction fragments length polymorphisms and copy numbers, sixty overlapping fragments derived from the 260-kb contig were used as probes to compare their corresponding structures among the Oryza species with AA-genome. According to the hybridization patterns, each fragment was classified into four types; true single copy (class 1), single copy with a smear background (class 2), multiple copy without a smear background (class 3), and only a smear background (class 4). Out of 16 single copy (class 1 and class 2) regions obtained in this map, the one site corresponding to wx gave rise to remarkable polymorphisms among AA-genome species in Oryza. In most of the fragments observed as repetitive segments (class 4), we could not find obvious differences in the hybridization pattern. However, interestingly, one site sorted into class-3 showed copy numbers varying among the lines. The lines belonging to O. sativa O. rufipogon, O. meridionalis,and O. longistaminata possessed high-copy numbers of this fragment, whereas only a few bands were detected in the lines from O. glaberrima, O. barthii, and O. glumaepatula. The two variable regions found within the AA-genome species represented genomic dynamisms. Received: 3 February 1999 / Accepted: 22 June 1999     

Microsatellites and microsynteny in the chloroplast genomes of Oryza and eight other Gramineae species

Primer pairs flanking ten chloroplast microsatellite loci, originally identified in Oryza sativa cv Nipponbare, were evaluated for amplification and allelic diversity using a panel of 13 diverse cultivars of rice (O. sativa), 19 accessions of wild rice (three O. officinalis, five O. latifolia, five O. minuta, four O. australiensis, one O. brachyantha and one O. ridleyi) and eight other Gramineae species (maize, teosinte, wheat, oat, barley, pearl millet, sorghum and sugarcane). Amplified products were obtained for all samples at nine out of ten loci. Among the rice cultivars, the number of alleles per locus ranged from one to four, with monomorphic patterns observed at five loci. The average polymorphism information content (PIC) value at the other five (polymorphic) loci was 0.54 among the 13 cultivars. When wild rice and the other Gramineae species were compared based on the proportion of shared alleles, their phylogenetic relationships were in agreement with previous studies using different types of markers; however, the magnitude of the differences based on chloroplast microsatellites underestimated the genetic distance separating these divergent species and genera. A sequence-based comparison of homologous regions of the rice and maize chloroplast genomes revealed that, while a high level of microsynteny is evident, the occurrence of actively evolving microsatellite motifs in specific regions of the rice chloroplast genome appears to be mainly a species or genome-specific phenomenon. Thus the chloroplast primer pairs used in this study bracketed mutationally active microsatellite motifs in rice but degenerate, interrupted motifs or highly conserved, mutationally inert motifs in distantly related genera. Received: 17 March 1999 / Accepted: 11 November 1999     

Direct amplification of minisatellite-region DNA with VNTR core sequences in the genus Oryza

 A polymerase chain reaction (PCR) application, involving the directed amplification of minisatellite-region DNA (DAMD) with several minisatellite core sequences as primers, was used to detect genetic variation in 17 species of the genus Oryza and several rice cultivars (O. sativa L.). The electrophoretic analysis of DAMD-PCR products showed high levels of variation between different species and little variation between different cultivars of O. sativa. Polymorphisms were also found between accessions within a species, and between individual plants within an accession of several wild species. The DAMD-PCR yielded genome-specific banding patterns for the species studied. Several DAMD-PCR-generated DNA fragments were cloned and characterized. One clone was capable of detecting multiple fragments and revealed individual-specific hybridization banding patterns using genomic DNA from wild species as well as rice cultivars. A second clone detected only a single polymorphic locus, while a third clone expressed a strong genome specificity by Southern analysis. The results demonstrated that DAMD-PCR is potentially useful for species and genome identification in Oryza. The DAMD-PCR technique also allows for the isolation of informative molecular probes to be utilized in DNA fingerprinting and genome identification in rice. Received: 1 October 1996 / Accepted: 25 April 1997     

Geographic distribution and multilocus organization of isozyme variation of rice (Oryza sativa L.)

Genetic organization of isozyme variation in rice (Oryza sativa L.) was investigated based on 17 polymorphic isozyme loci using a sample of 511 accessions of worldwide origin. The genetic diversity within the species was very high (H=0.36 with 4.82 alleles per locus), as compared with most selfing plant species. Three diversity centers were detected for isozyme variation including South Asia, China and Southeast Asia. The accessions were classified into three well-differentiated cultivar groups corresponding to the indica and japonica subspecies, and a new unnamed group. Variation within the cultivar groups accounted for 80% of the total isozyme variation. Within-country variation accounted for 58% of the total variation while among-region and among-country variation within the cultivar groups accounted for only 14% and 8% of the total variation. Analyses using log-linear models revealed that pronounced non-random associations between and among alleles at many unlinked isozyme loci were organized in a non-hierarchical pattern, and subspecific and macro-geographic differentiation was much more pronounced in multilocus phenotype frequencies than in allelic frequencies at individual loci. These results suggest that selection on multilocus gene complexes was largely responsible for the maintenance of the extensive isozyme variation within the species and the indica-japonica differentiation. Our results further suggest the independent domestication of indica and japonica, the dual origins of the indica rice from China and South Asia (India), and the differentiation of the ecotypes ’javanica’ and the ’temperate japonica’ within the japonica subspecies. Received: 5 August 1999 / Accepted: 13 December 1999     

Genetic diversity and phylogenetic relationship as revealed by inter simple sequence repeat (ISSR) polymorphism in the genus Oryza

Inter simple sequence repeat (ISSR) polymorphism was used to determine genetic diversity and phylogenetic relationships in Oryza. Forty two genotypes including 17 wild species, representing AA,BB,CC,EE,FF,GG,BBCC,CCDD, and HHJJgenomes, two cultivated species, Oryza sativa (AA) and Oryza glaberrima (AA), and three related genera, Porteresia coarctata, Leersia and Rhynchoryza subulata, were used in ISSR analysis. A total of 30 ISSR primers were screened representing di-, tri-, tetra- and penta-nucleotide repeats, of which 11 polymorphic and informative patterns were selected to determine the genetic diversity. The consensus tree constructed using binary data from banding patterns generated by ISSR-PCR clustered 42 genotypes according to their respective genomes. ISSR analysis suggests that the genus Oryza may have evolved following a polyphyletic pathway; Oryza brachyantha (FF genome) is the most divergent species in Oryza and Oryza australiensis (EE genome) does not fall under the Officinalis complex. DNA profiles based on ISSR markers have revealed potential diagnostic fingerprints for various species and genomes, and also for individual accessions/cultivars. Additionally ISSR revealed 87 putative genome/species-specific molecular markers for eight of the nine genomes of Oryza. The ISSR markers are thus useful in the fingerprinting of cultivated and wild species germplasm, and in understanding the evolutionary relationships of Oryza. Received: 23 August 1999 / Accepted: 10 November 1999     

PCR-based molecular markers for the fragrance gene in rice (Oryza sativa. L.)

The genomic DNA clone RG28, linked to the major fragrance gene of rice (fgr), was assessed for polymorphism in order to produce a PCR-based marker for fragrance. A small mono-nucleotide repeat, that was polymorphic between a pair of fragrant and non-fragrant cultivars, was identified and developed into a co-dominant PCR-based marker. The polymorphism-information-content determinations for three microsatellite markers, that have been genetically mapped near RG28, are also presented. These PCR-based markers will be highly useful in distinguishing fragrance-producing alleles from non-fragrance-producing alleles at the fgr locus. Received: 19 October 1999 / Accepted: 16 December 1999     

The development of oat microsatellite markers and their use in identifying relationships among Avena species and oat cultivars

Microsatellites have many desirable marker properties. There has been no report of the development and utilization of microsatellite markers in oat. The objectives of the present study were to construct oat microsatellite-enriched libraries, to isolate microsatellite sequences and evaluate their level of polymorphism in Avena species and oat cultivars. One hundred clones were isolated and sequenced from three oat microsatellite-libraries enriched for either (AC/TG) _n , (AG/TC) _n or (AAG/TTC) _n repeats. Seventy eight clones contained microsatellites. A database search showed that 42% of the microsatellite flanking sequences shared significant homology with various repetitive elements. Alu and retrotransposon sequences were the two largest groups associated with the microsatellites. Forty four primer sets were used to amplify the DNA from 12 Avena species and 20 Avena sativa cultivars. Sixty two percent of the primers revealed polymorphism among the Avena species, but only 36% among the cultivars. In the cultivars, the microsatellites associated with repetitive elements were less polymorphic than those not associated with repetitive elements. Only 25% of the microsatellites associated with repetitive elements were polymorphic, while 46% of the microsatellites not associated with repetitive elements showed polymorphism in the cultivars. An average of four alleles with a polymorphism information content (PIC) of 0.57 per primer set was detected among the Avena species, and 3.8 alleles with a PIC of 0.55 among the cultivars. In addition, 54 barley microsatellite primers were tested in Avena species and 26% of the primers amplified microsatellites from oat. Using microsatellite polymorphisms, dendrograms were constructed showing phylogenetic relationships among Avena species and genetic relationships among oat cultivars. Received: 1 November 1999 / Accepted: 14 April 2000     

Characteristics of genetic variation in the progenies of protoplast-derived plants of rice, Oryza sativa cv Nipponbare

Genetic variation in protoplast-derived rice (Oryza sativa L.) plants was characterized using first and second generation selfed progenies. A total of 133 regenerated plants were obtained from ten protoplasts of the japonica rice cultivar Nipponbare. Sixty two regenerated plants which set enough seeds for the subsequent field tests at the next generation and were derived from five protoplasts were selected, and their selfed seeds were used as the first selfed-seed progeny generation). Fifteen plants were selected from each of the 15 lines, and their selfed seeds were used for tests at the generation. Thirty seven lines (60%) segregated plants with detrimental mutant characters of yellow-green phenotype, dwarf stature, dense and short panicle, or low seed fertility. According to the segregation patterns in the lines having mutated plants among those originated from the same protoplasts, the stages of mutation induction were estimated. Additionally, five quantitative traits were changed in almost all and lines. Varied quantitative traits of heading date, number of spikelets per panicle, and seed fertility, were in a heterozygous state. However, culm and panicle lengths showed high uniformity, whereas reduced culm and panicle lengths were caused by mutational changes in polygenes and/or multiple genes. Received: 20 March 1996 / Accepted: 21 June 1996     

Phylogenetic relationships among A-genome species of the genus Oryza revealed by intron sequences of four nuclear genes

The A-genome group in Oryza consists of eight diploid species and is distributed world-wide. Here we reconstructed the phylogeny among the A-genome species based on sequences of nuclear genes and MITE (miniature inverted-repeat transposable elements) insertions. Thirty-seven accessions representing two cultivated and six wild species from the A-genome group were sampled. Introns of four nuclear single-copy genes on different chromosomes were sequenced and analysed by both maximum parsimony (MP) and Bayesian inference methods. All the species except for Oryza rufipogon and Oryza nivara formed a monophyletic group and the Australian endemic Oryza meridionalis was the earliest divergent lineage. Two subspecies of Oryza sativa (ssp. indica and ssp. japonica) formed two separate monophyletic groups, suggestive of their polyphyletic origin. Based on molecular clock approach, we estimated that the divergence of the A-genome group occurred c. 2.0 million years ago (mya) while the two subspecies (indica and japonica) separated c. 0.4 mya. Intron sequences of nuclear genes provide sufficient resolution and are informative for phylogenetic inference at lower taxonomic levels.     

水稻CMS相关基因在稻属AA基因组中的分布及其单核苷酸多态性

水稻线粒体基因组嵌合基因orf79 和 orfH79分别被认为与BT-型和HL-型水稻CMS有关, 两者具有98%的同源性, 并且其DNA序列只存在4核苷酸的差异。对于这两个嵌合基因, 前者来源于栽培稻(Oryza. sativa L.), 而后者则来源于普通野生稻(O. rufipogon Griff.)。这意味着orf79/ orfH79可能在广泛分布于稻属AA基因组中。为了调查orf79/ orfH79在稻属物种中的分布和变异, 190份栽培稻品系[包括156份亚洲栽培稻(O. sativa var. landrace)和34份非洲栽培稻(O. glaberrima)]以及104份稻属AA基因组野生稻品系(包括O. rufipogon、O.nivara、O. glumaepatula、O. barthii、O. longistaminata和O. meridionalis 6个种), 被用于PCR扩增检测。31份具有控制粤泰A和笹锦A的特异片段的稻属AA基因组水稻品系被检测出。所有特异片段均被回收并测序, 基于DNA 序列的聚类结果显示31份水稻材料被分成了两组, 分别代表为BT-型和HL-型水稻不育细胞质组群。结果也进一步表明: HL-型水稻CMS胞质主要分布于一年生的O. nivara中; BT-型水稻CMS胞质可能来源于栽培稻变种或多年生野生稻O. rufipogon。     

Polymorphism and phylogenetic relationships among species in the genus Oryza as determined by analysis of nuclear RFLPs

Summary Ninety-three accessions representing 21 species from the genus Oryza were examined for restriction fragment length polymorphism. The majority (78%) of the accessions, for which five individuals were tested, were found to be monomorphic. Most of the polymorphic accessions segregated for only one or two probes and appeared to be mixed pure lines. For most of the Oryza species tested, the majority of the genetic variation (83%) was found between accessions from different species with only 17% between accessions within species. Tetraploid species were found to have, on average, nearly 50% more alleles (unique fragments) per individual than diploid species reflecting the allopolyploid nature of their genomes.Classification of Oryza species based on RFLPs matches remarkably well previous classifications based on morphology, hybridization and isozymes. In the current study, four species complexes could be identified corresponding to those proposed by Vaughan (1989): the O. ridleyi complex, the O. meyeriana complex, the O. officinalis complex and the O. sativa complex. Within the O. sativa complex, accessions of O. rufipogon from Asia (including O. nivara) and perennial forms of O. rufipogon from Australia clustered together with accessions of cultivated rice O. sativa. Surprisingly, indica and japonica (the two major subspecies of cultivated rice) showed closer affinity with different accessions of wild O. Rufipogon than to each other, supporting a hypothesis of independent domestication events for these two types of rice. Australian annual wild rice O. meridionalis (previously classified as O. rufipogon) was clearly distinct from all other O. rufipogon accessions supporting its recent reclassification as O. meridionalis (Ng et al. 1981). Using genetic relatedness as a criterion, it was possible to identify the closest living diploid relatives of the currently known tetraploid rice species. Results from these analyses suggest that BBCC tetraploids (O. malampuzhaensis, O. punctata and O. minuta) are either of independent origins or have experienced introgression from sympatric C-genome diploid rice species. CCDD tetraploid species from America (O. latifolia, O. alta and O. grandiglumis) may be of ancient origin since they show a closer affinity to each other than to any known diploid species. Their closest living diploid relatives belong to C genome (O. eichingeri) and E genome (O. Australiensis) species. Comparisons among African, Australian and Asian rice species suggest that Oryza species in Africa and Australia are of polyphyletic origin and probably migrated to these regions at different times in the past.Finally, on a practical note, the majority of probes used in this study detected polymorphism between cultivated rice and its wild relatives. Hence, RFLP markers and maps based on such markers are likely to be very useful in monitoring and aiding introgression of genes from wild rice into modern cultivars.     

Genetic relationships among Stylosanthes species revealed by RFLP and STS analyses

Genetic relationships between 6 unclassified taxa and 24 known species of the genus Stylosanthes were investigated by RFLP and STS analyses. This allowed the diploid taxa used in this study to be classified into nine basal (genome) groups. Representative species in these groups included ’S. seabrana’/S. hamata (Group A), S. viscosa (Group B), S. humilis (Group C), S. macrocephala/S. bracteata (Group D), S. pilosa (Group E), S. leiocarpa (Group F), S. guianensis (Group G), S. tomentosa (Group H) and S. calcicola (Group I). Polyploid taxa used were grouped into five classes based on their putative genomic structures. These are AABB for S. scabra, S. aff. scabra, S. sericeiceps, S. aff. hamata and S. tuberculata; AACC for S. mexicana, S. subsericea, S. sundaica and S. sp.A; DDEE for S. capitata; AAFF for S. sympodialis; and AABBXX for S. erecta, with XX representing an unknown genome. Of the 6 unclassified taxa, three were diploids and 3 tetraploids. Of the 3 diploids, the genome of S. sp. was markedly distinct from those of all other diploids analysed in this study, with that of S. leiocarpa being the closest. The genome of S. sp.B was similar to that of S. humilis, with an average dissimilarity value of 15% between them. The genome of S. aff. viscosa was very similar to that of S. viscosa. Genetic variation between these 2 taxa was not larger than that within each of the 2 taxa. Of the 3 tetraploids, the genomic structure of S. sp.A was similar to those of S. mexicana, S. sundaica and S. subsericea, and the genomic structures of S. aff. scabra and S. aff. hamata were similar to those of S. scabra and S. sericeiceps. Received: 18 September 1998 / Accepted: 23 March 1999     

Isolation and characterization of OsDMC1, the rice homologue of the yeast DMC1 gene essential for meiosis

Yeast DMC1 is a meiosis-specific gene required for homologous chromosome pairing in meiosis. Using degenerate primers designed according to amino acid motifs conserved in yeast Dmc1 and Arabidopsis AtDmc1, we obtained full-length cDNA of a rice homologue of the DMC1 gene (OsDMC1) by RT-PCR and rapid amplification of cDNA ends (RACEs). OsDmc1 exhibited 53% amino acid sequence identity to yeast Dmc1 and 81% to AtDmc1. OsDMC1 was expressed at high-levels in reproductive organs, low-levels in roots, and undetectable levels in leaves and seedlings. Southern blot analyses revealed that OsDMC1 is one of two DMC1 homologues present in rice. Received: 18 December 2000 / Accepted: 22 December 2000     

A combined RFLP and AFLP linkage map of upland rice (Oryza sativa L.) used to identify QTLs for root-penetration ability

Acombined RFLP and AFLP linkage map of an F₆ recombinant inbred population, which was derived from a previously mapped F₂ of a cross between the two drought resistant upland rice varieties Bala and Azucena, is presented. The map contains 101 RFLP and 34 AFLP markers on 17 linkage groups covering 1680 cM. Also presented is the approximate mapping position of a further four RFLP and 75 AFLP markers, which either could not be given a unique place on the map or for which the available data is not sufficient to allow confident positioning, and the result of quantitative trait locus (QTL) mapping of traits related to root-penetration ability. Root penetration was assessed by counting the number of root axes that penetrated a 3 mm-thick layer consisting of 80% wax and 20% white soft paraffin. Good root penetration would be expected to increase drought resistance where soil strength is high. Single-marker analysis revealed seven QTLs for the number of roots which penetrate the wax layer. In identical locations were seven QTLs for the ratio of penetrated to the total number of roots. Transgressive inheritance of positive alleles from Bala explained four of these QTLs. Comparison of the QTLs identified here with previous reports of QTLs for root morphology suggest that alleles which improve root penetration ability may also either make the roots longer or thicker. Received: 3 February 1999 / Accepted: 30 April 1999     

Evolutionary relationship of plant catalase genes inferred from exon-intron structures: isozyme divergence after the separation of monocots and dicots

In order to understand the molecular evolution of catalase genes in higher plants, we compared the exon-intron structures of 12 genomic sequences from six plant species. It was assumed that the putative single primordial catalase gene had seven introns, because only those catalase genes having this structure are found in the monocotyledonae and dicotyledonae classes. After the evolutionary divergence of monocots from dicots, consecutive duplication of the primordial gene followed by the differential loss of introns occurred in each class to form three (or possibly four in dicots) diverse isozyme genes. In monocots, three ancestral isozyme genes were formed before the divergence of ancestral rice and maize. One of the rice genes, CatA, has an entirely new short intron which was not found in any other plant catalase gene examined. We have investigated the existence of the intron in the CatA homolog in other rice species by polymerase chain reaction (PCR) analysis. One major PCR product was found with the genomic DNAs from O. sativa (indica and japonica types), O. rufipogon and O. glaberrima. DNAs from several accessions of O. longistaminata showed variation in both the number and size of the DNA fragments amplified. PCR analyses and sequencing of the PCR products revealed that there are several CatA homologs having different sequences in some accessions of O. longistaminata. We have extended our study to other species in the Poaceae. The results suggest that the gain of the intron, most likely by insertion of a retroposon, took place in the ancestral genome of rice after its evolutionary divergence from other ancestral cereals such as barley, wheat and oat. Received: 20 November 1997 / Accepted: 5 January 1998     

Inter- and intra-specific distribution of Stowaway transposable elements in AA-genome species of wild rice

The presence or absence of miniature inverted-repeat transposable elements (MITEs) that belong to Stowaway family was analyzed at three loci, two of which are newly identified, in five wild rice species having the AA genome. The pattern of the presence or absence of MITEs was found to be highly associated with speciation in this plant group. In Oryza rufipogon, the pattern was also associated with differentiation into annual or perennial ecotypes. These results suggest either that gene flow has been highly restricted between different species, as well as between different ecotypes of O. rufipogon after they were differentiated, or that loci with or without MITEs have been selected in nature together with the linked genes that are responsible for adaptation to environments. In addition, a very low polymorphism with regard to the presence or absence of MITEs within each species or each ecotype suggests that the frequency of transposition of MITEs is very low, assuming that the loci that contain MITEs are free from selection pressure. Received: 30 October 1999 / Accepted: 2 December 1999     

Genetic variation within and among populations of a wild rice Oryza granulata from China detected by RAPD and ISSR markers

Genetic variation within and between five populations of Oryza granulata from two regions of China was investigated using RAPD (random amplified polymorphic DNA) and ISSR (inter-simple sequence repeat amplification) markers. Twenty RAPD primers used in this study amplified 199 reproducible bands with 61 (30.65%) polymorphic; and 12 ISSR primers amplified 113 bands with 52 (46.02%) polymorphic. Both RAPD and ISSR analyses revealed a low level of genetic diversity in wild populations of O. granulata. Furthermore, analysis of molecular variance (AMOVA) was used to apportion the variation within and between populations both within and between regions. As the RAPD markers revealed, 73.85% of the total genetic diversity resided between the two regions, whereas only 19.45% and 6.70% were present between populations within regions and within a population respectively. Similarly, it was shown by ISSR markers that a great amount of variation (49.26%) occurred between the two regions, with only 38.07% and 12.66% between populations within regions and within a population respectively. Both the results of a UPGMA cluster, based on Jaccard coefficients, and pairwise distance analysis agree with that of the AMOVA partition. This is the first report of the partitioning of genetic variability within and among populations of O. granulata at the DNA level, which is in general agreement with a recent study on the same species in China using allozyme analysis. Our results also indicated that the percentage of polymorphic bands (PPB) detected by ISSR is higher than that detected by RAPD. It seems that ISSR is superior to RAPD in terms of the polymorphism detected and the amplification reproducibility. Received: 29 March 2000 / Accepted: 15 May 2000