Genetic diversity of Pakistan wheat germplasm as revealed by RAPD markers

Wheat breeding in Pakistan started in 1930s before partition in the United India and so far has released more than 68 cultivars, but no systematic analyses of the genetic diversity of Pakistan wheat have been made. Twenty Pakistan wheat cultivars released from 1933 to 2002 were examined for genetic diversity and relationships using random amplified polymorphic DNA (RAPD) markers. Forty-two RAPD primers were applied and 184 polymorphic bands were generated for each cultivar. Most of the cultivars were genetically interrelated, although six of them displayed some genetic distinctness. The RAPD variation observed among these cultivars was low. Only 40.7% of the total scorable bands were polymorphic, and 26.1% of the polymorphic bands were observed most frequently (f = 0.95) among the 20 cultivars. The proportions of polymorphic bands for each cultivar ranged from 0.67 in ‘Yecora’ to 0.84 in ‘C-250’ with an average of 0.76. About 1.4% of the RAPD variation might have been fixed over the 69 years of wheat breeding, but such fixation was not statistically significant. These results are significant for future improvement and conservation of Pakistan wheat.     

Genetic diversity among <Emphasis Type="Italic">Jatropha</Emphasis> species as revealed by RAPD markers

The genus Jatropha is native of tropical America with more than 200 species that are widely distributed in tropics with a promise for use as an oil crop for biodiesel. This investigation was carried out to assess the genetic diversity of 12 Jatropha species based on random amplified polymorphic DNA markers. From 26 random primers used, 18 primers gave reproducible amplification banding patterns of 112 polymorphic bands out of 134 bands scored accounting for 80.2% polymorphism across the genotypes. Three primers viz., OPA 4, OPF 11, and OPD 14 generated 100% polymorphic patterns. The polymorphic information content was highest for the primer OPD 14 (0.50) followed by the primers OPF 11 and OPAD 11 (0.48). Jaccard’s coefficient of similarity varied from 0.00 to 0.85, indicative of high level of genetic variation among the genotypes studied. UPGMA cluster analysis indicated three distinct clusters, one comprising all accessions of J. curcas L., while second included six species viz., J. ramanadensis Ramam., J. gossypiifolia L., J. podagrica Hook., J. tanjorensis J. L. Ellis et Saroja J. villosa Wight and J. integerrima Jacq. J. glandulifera Roxb. remained distinct and formed third cluster indicating its higher genetic distinctness from other species. The overall grouping pattern of clustering corresponds well with principal component analysis confirming patterns of genetic diversity observed among the species. The result provides valid guidelines for collection, conservation and characterization of Jatropha genetic resources.     

Genetic diversity and differentiation in Ethiopian populations of Phytolacca dodecandra as revealed by AFLP and RAPD analyses

Amplified fragment length polymorphism (AFLP) and random amplified polymorphic DNA (RAPD) analyses were performed on six populations (a total of 89 individuals) of Phytolacca dodecandra (endod) collected in Ethiopia. The populations were selected based on our previous investigation to represent two altitude groups: lowland/central-highland (1600–2500 m) and highland (2501–3000 m). A total of 197 AFLP and 68 RAPD markers were scored from 5 primer pairs and 12 random primers, respectively. The overall patterns obtained for AFLPs and RAPDs from diversity, cluster and principal component analyses were very comparable. However, the moderate correlation (r = 0.56) between AFLP and RAPD similarity matrices as well as the discrepancies in diversity estimates between the two techniques in some populations and in the lowland/central-highland plants could be due to differences in sensitivity of reaction conditions, bias in scoring of bands, number of markers used for analyses, and/or parts of the genome surveyed. For both AFLP and RAPD, the lowland/central-highland populations showed higher polymorphism and Shannon information measure (H) than the highlands. Cluster and principal component analyses performed for both marker types have also clearly demonstrated the differentiation of all the lowland/central highland plants from those of the highlands, in agreement with our previous conclusion. Markers scored from any of the five AFLP primer pairs were sufficient to clearly distinguish the two altitude groups; with RAPD, selection of about 8 informative markers produced by seven random primers was needed for the same purpose.     

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.     

Genetic structure of quinoa (<Emphasis Type="Italic">Chenopodium quinoa</Emphasis> Willd.) from the Bolivian altiplano as revealed by RAPD markers

Quinoa (Chenopodium quinoa Willd.) is a pseudocereal originated from the Andes important for small farmers’ food security as well as for commercial production. Recently, it has been claimed that in Bolivia genetic erosion could result from the marginalization of the crop in the north and from its commercial standardization in the south. The aim of this study was to quantify the hierarchical structure of the genetic variation present in eight quinoa field populations, consisting of cultivated and weedy individuals, representative of the altiplano and interandean valleys of Bolivia. Randomly amplified polymorphic DNA markers show that quinoa has a strong population structure and a high intra-population variation. An effect of geographical structure of the populations was highlighted, due to population isolation, not simply linked to distance but more probably to climatic and orographic barriers present in the studied zone. The population structure is also reinforced by the limited seed exchanges among farmers as revealed by field interviews. This population structure appears related to three major biogeographic zones: the northern and central altiplano, the interandean valley, and the southern Salar. Intrapopulation genetic diversity was higher than that expected for a mainly autogamous species, and higher than that reported in anterior studies based on germplasm collections. These results are commented in view of current knowledge on phylogeny and reproductive biology of the species, and their implications regarding genetic resources management are discussed.     

Genetic diversity and relationships among cultivated and wild accessions of tartary buckwheat (Fagopyrum tataricum Gaertn.) as revealed by RAPD markers

The methodology of sampling and the selection of a proper marker systemfor the analysis of accessions are major concerns in the evaluation of gene bank material. In our study the RAPD analysis of bulked DNA samples and single seedsDNA was successfully employed to evaluate intra- and inter-population geneticvariability of cultivated and wild tartary buckwheat accessions. The bulkingapproach enabled the distinction of all 40 analysed accessions and theirseparation into geographically well defined clusters. Three wild populations,two from Sichuan and one from Qinghai, formed a group that was geneticallyrelatively distant from wild populations from Tibet and all cultivatedlandraces which, on the other hand, exhibited very close relationships. Thesingle seed study that was used after bulked DNA analysis provided detailedinformation of the genetic variation present within some accessions of specialinterest. A moderate level of genetic variability was detected betweenaccessions and the variability was partitioned into between- andwithin-population components. On average, most of the detected variation ispresent between F. tataricumpopulations. The genetic and geographic distribution of variability is furtherdiscussed. We demonstrated the usefulness of combining bulking and single seedstudy approaches for the effective evaluation of genetic variability inF. tataricum accessions that couldalso have wider applicability in the management of plant genetic resources andphylogenetic studies.     

Genetic diversity of <Emphasis Type="Italic">Nelumbo</Emphasis> accessions revealed by RAPD

Total 65 lotus accessions in genus Nelumbo mainly collected from China, were subjected to random amplified polymorphic DNA (RAPD) markers to estimate the genetic diversity and to test the genetic basis of the relationships between morphotypes and molecular markers. Seventeen primers generated a total of 195 highly reproducible and discernible loci, among which 173 were polymorphic. Percent polymorphism varied from 66.7 to 100 with an average of 88.72, and five primers out of them, OPC05, OPG10, OPN20, OPP09 and OPS17, showed 100% polymorphism. A relatively high genetic diversity was detected among all the samples with the similarity coefficient values ranging from 0.45 to 0.85, and Nei’s gene diversity (h) 0.30, and Shannon index (I) 0.46. The UPGMA dendrogram clustered 65 accessions in four clusters and the clustering pattern showed two groups, N. nucifera ssp. nucifera and those accessions related to the American lotus, and some special cultivars, landraces, hybrids and the American lotus. Principal Coordinate Analysis (PCA) further indicated that the genetic diversity of Nelumbo accessions was not evenly distributed, instead, was presented by a clustered distribution pattern. Similar to the results revealed by the dendrogram, two main groups representing the two subspecies of N. nucifera, as well as some special landraces, cultivars of Chinese lotus, the Japanese lotus and hybrids out of the two groups were obtained. Neither the UPGMA dendrogram nor the PCA analysis exhibited strict relationship with geographic distribution and morphotypes among the accessions.     

The Vigna angularis complex: Genetic variation and relationships revealed by RAPD analysis, and their implications for in situ conservation and domestication

The present study, using RAPD analysis, was undertaken to characterize genetic variation in three forms of V. angularis, cultivated, wild and weedy forms, and their relationships. The materials used consisted of 171 individuals (plants) or cultivars from 23 populations including 5 wild populations, 6 weedy populations, 6 cultivated populations and 6 populations with plants having wild and weedy or intermediate morphology, denoted here as complex populations. The materials used were collected on Honshu Island, Japan and seeds collected directly from the field were germinated for DNA extraction. In addition, 6 landrace accessions of V. angularis from the genebank were also analyzed. Genetic variation was highest in the wild form (H_g= 0.132; GD = 0.388), followed by the weedy form (H_g= 0.124; GD = 0.341) and the least in the cultivated form (H_g= 0.079; GD = 0.274). Intra-population genetic variation was high in the weedy and in the wild populations. However, inter-population was greater than intra-population genetic variation for all groups of populations studied in the V. angularis complex. 93% of the total diversity in the present study was exhibited by plants from complex populations and specific RAPD bands were found in these populations. Our results provide evidence that complex populations would be a logical focus for efforts to conserve the V. angularis complex in situ. Our results suggest that weedy populations are usually an ecotype of the wild form adapted to a different habitat.     

Genetic diversity of Central Asian and north Caucasian Aegilops species as revealed by RAPD markers

RAPD analysis of 112 accessions of Aegilops tauschii Coss. (genome DD), Ae. cylindrica Host (CCDD), Ae. crassa Boiss. (DDMM), Ae. biuncialis Vis. (UUMM) and Ae. triuncialis L. (UUCC) collected in the Central Asia and north Caucasia was conducted. Aegilops accessions were divided into two major groups, corresponding to the D genome species and the U genome species. These groups were also separated into sub-groups according to species, except for the Ae. tauschii-cylindrica complex of accessions from Central Asia. Aegilops tauschii from north Caucasia was divided into two varietal groups, tauschii and meyeri. The Central Asian accessions of Aegilops species were more diverse than the accessions from north Caucasia. Aegilops tauschii and Ae. cylindrica accessions from north Caucasia were genetically uniform. Associations between altitudal variation of Aegilops species and variability of RAPD markers were not found.     

Genetic Diversity of Traditional Chinese Mustard Crops Brassica juncea as Revealed by Phenotypic differences and RAPD Markers

This investigation was aimed at exploring the genetic diversity among nine typical accessions of Chinese mustard crops using random amplified polymorphic DNA (RAPD) markers and morphological comparison. Totally, 111 reproducible DNA bands were generated by 16 arbitrary primers, of which 91 bands were proved to be polymorphic. Based on pair-wise comparisons of the amplified bands, genetic similarities were obtained using Nei & Li's similarity coefficients and a dendrogram reflecting their relationships was made using the unweighted pair–group method with arithmetic averages (UPGMA). The result of cluster analysis indicated that the nine accessions were capable of being classified into two primary groups, one including accession 2 with expanded root (root mustard), accession 3 with entirely expanded whole stem (long-stem mustard), accession 6 with edible leaves (leaf mustard), accession 8 with edible seed stalk (seed stalk mustard) and another one including accession 4 with expanded basal stem (short-stem mustard), accession 5 with bulgy petiole (leafy bulgy mustard), and accession 9 with mustard-rich seed (seed mustard). Besides, accession 1 with expanded root (root mustard) and accession 7 with edible leaves and seed stalk (seed stalk mustard) were independent of other accessions in the dendrogram. Additionally, by cluster analysis based on highly reproducible RAPD markers, the accessions with similar edible parts of leaves or roots were not actually in the same phylogenetic groups. This implied that they were probably derived from different geographical origins with dissimilar genetic background and possessed higher genetic diversification. Furthermore, the results indicated that the traditional method for classifying Chinese mustard crops was not much reliable as it was largely dependent on phenotypic behaviors. Meanwhile, the phenotypic differences among individuals did not necessarily mean they must have sharp difference in genetic background as they met in the same group. Undoubtedly, these results aforementioned make this crop quite interesting to researchers for further investigating the molecular evolution of this special AABB group.     

Genetic variation in captive population of chinese alligator, Alligator sinensis, revealed by random amplified polymorphic DNA (RAPD)

Chinese alligator (Alligator sinensis) is an endemic species in China. The likely extinction of it in the wild has been recognised. To prevent this species becoming extinct, the Anhui Research Centre of Chinese Alligator Reproduction (ARCCAR) was established in Xuanzhou, Anhui Province in 1979, where has been established the largest captive population of Chinese alligator (XZSP) in the world. Another farm (CXSP) was established by villagers in Changxin, Zhejiang Province. The results of an investigation of the two captive subpopulation structures by genetic analysis are presented in this paper. We examined the genetic variation in the two captive subpopulations using RAPDs. Thirty-one random primers were selected among 199 random primers screened. A total of 193 reproducible RAPD fragments were scored among 43 individuals, of which 21 (10.88%) were polymorphic. The genetic distances between 43 individuals ranged from 0 to 0.0376 with average of 0.0104±0.0055 S.E. The genetic similarity in CXSP (0.9948±0.0029 S.E.) was higher than that in XZSP (0.9894±0.0055 S.E.). The founder effect is a possible explanation for very low genetic variation in CXSP. Analysis of the RAPD data showed that the mean phenotypic band frequencies of each polymorphic loci was 0.6656±0.3730 S.E. The lowest phenotypic band frequency (0.0233) was found in four of those polymorphic loci. There was no genetic difference between the two subpopulations (D_ij=0.0009). According to the dendrogram and the distribution of polymorphic fragments in two subpopulations, CXSP originated genetically from XZSP. This paper summarises a preliminary research on genetic structure in populations of Chinese alligator. Although there is higher genetic similary (0.9896±0.0055 S.E.) in captive population of A. sinensis, we did not determine whether or not loss of genetic variation had occurred in relation to a wild control population. The data of malformed offspring will be collected carefully, and wild samples be added to set up a control population in future study.     

Genetic diversity of <Emphasis Type="Italic">Guizotia abyssinica</Emphasis> (L. f.) Cass. (Asteraceae) from Ethiopia as revealed by random amplified polymorphic DNA (RAPD)

Genetic diversity of 70 populations of niger (Guizotia abyssinica) representing all its growing regions in Ethiopia was investigated using random amplified polymorphic DNA (RAPD) to reveal the extent of its populations genetic diversity. Ninety-seven percent of the loci studied was revealed to be polymorphic for the whole data set. The within population diversity estimated by Shannon diversity index and Nei gene diversity estimates was revealed to be 0.395 and 0.158, respectively. The extent of genetic variation of populations from major niger producing regions was significantly lower than that of populations from other regions; however, it is distributed regardless of altitude of growth. Genetic differentiation between populations was estimated with Shannon index as G^′ _ST (0.432), Nei’s G _ST (0.242) and AMOVA based F _ST (0.350) and appears to be equivalent to the average values calculated from various RAPD based studies on outcrossing species. Higher proportion of the variation detected by AMOVA resided within populations (64.58%) relative to the amount of variation among populations (35.42%). UPGMA cluster analysis showed that most of the populations were clustered according to their region of origin. However, some populations were genetically distant from the majority and seem to have unique genetic properties. It is concluded that the crop has a wide genetic basis that may be used for the improvement of the species through conventional breeding and/or marker assisted selection. Collection of germplasm from areas not yet covered and/or underrepresented is the opportunity to broaden the genetic basis of genebank collection.     

Genetic diversity in the Olive tree (Olea europaea L. subsp. europaea) cultivated in Portugal revealed by RAPD and ISSR markers

To assess the genetic diversity of the most important olive cultivars used in Portugal, a base collection was established with two hundred and one accessions of eleven cultivars from the different agro-ecological-regions (AER) of olive oil production. Inter-cultivar diversity was evaluated using seven RAPD primers producing fifty-nine polymorphic markers that enable cultivar distinction. Discriminant analysis according to fruit use and AER revealed a genetic structure associated with local selection both for fruit exploitation and agro-ecological adaptation. Intra-cultivar diversity of the ancient cultivar Galega was also investigated. Three RAPD and five ISSR primers produced ninety-three polymorphic markers upon seventy-seven accessions from five AERs. Total accession discrimination was achieved. UPGMA clustering and discriminant analysis revealed that the genetic diversity was predominantly structured according to accessions origin. The within and among AER variation revealed by AMOVA supported this genetic structure and showed a high proportion of intra-AER variability. These evidences suggest that Galega is composed by a mixture of different genotypes adapted to local conditions, indicating that this cultivar is in an early stage of domestication and should be treated as a landrace instead of a uniform cultivar. The assessment of Galega genetic diversity within each of the five AERs indicated the highest significant level (H_g = 6.23 at p< 0.001) in Ribatejo-Santarém. This finding associated with the distinctiveness of Galega in relation to other Portuguese cultivars and with the recent insights of olive tree domestication allowed us to hypothesize that Ribatejo-Santarém was the ecological region of origin and dispersion of this ancient cultivar.     

Genetic relationships among perennial and annual Cicer species growing in Turkey as revealed by allozymes

Allozyme polymorphisms were used to assess genetic variation and relationships among ten Cicer species (annuals and perennials) growing in Turkey. Using seven enzyme systems, 12 putative scorable loci were detected and surveyed for polymorphism in an accession collection including wild and cultivated forms. Variation was generally low within accessions and species, but common between species. Cluster analysis based on the pairwise genetic distance coefficients among accessions and species using UPGMA revealed two species clusters; one includes three perennials (C. montbretii, C. isauricum and C. anatolicum) and the other contains six annuals (C. pinnatifidum, C. bijugum, C. judaicum, C. echinospermum, C. reticulatum and C. arietinum) and one perennial species (C.incisum). Grouping obtained in allozyme analysis appears to be consistent with the classification these species into three sections. However, contrary to relationships obtained in previous studies, three perennial species from section Polycicer were relatively distant to the group containing annuals. One perennial species, C. incisum from section Chamaecicer, clustered with annuals showing a close similarity. The grouping of six annual species was consistent with the previous reports of relationships. The relationships deduced between perennials and annuals appear to shed light on the evolution of annual habit from perennial habit.     

Genetic variations and interrelationships in Vanilla planifolia and few related species as expressed by RAPD polymorphism

Vanilla is naturally distributed in Mexico and parts of Central America and the history of origin of cultivated vanilla suggests that the entire stock outside Mexico may be from a single genetic source. In the present study, RAPD polymorphism was used to estimate the level of genetic diversity and interrelationships among different collections of Vanilla planifolia Andr., and few related species, including both leafy and leafless types such as V. tahitensis J.W.Moore, V. andamanica, Rolfe, V. pilifera Holtt., and V. aphylla Blume. Studies revealed that there are very limited variation within collections of V. planifolia, indicative of its narrow genetic base, and of the related species we tested, V. tahitensis is nearest to V. planifolia. The species studied are diverse and have a similarity ranging from 1.2 to 57.3 %. Of the sampled taxa, V. andamanica is the most divergent and there is also reasonable variability within its collections, indicating the possibility of natural seed set. A total of 82 polymorphic bands expressed in the RAPD profiles were used to generate a genetic distance matrix, which was then used in cluster analysis. Specific groupings were revealed by the cluster analysis whereby the leafless forms (V. aphylla, V. pilifera and the new species) and V. andamanica formed separate clusters. This is the first report of species interrelationship studies, including both cultivated and wild vanilla species.     

RAPD and seed coat morphology variation in annual and perennial species of the genus Cicer L.

Molecular and morphological variation of six perennial and five annual species including domesticated chickpea, C. arietinum, were inferred on the basis of RAPD and S.E.M. seed coat features using three outgroup taxa (Lens ervoides, Lathyrus japonica and Pisum sativum). Of the 66 decamer arbitrary primers tested, eight primers revealed 87 informative fragments. Neighbor-joining cluster analysis using Jaccard's coefficient of similarity on the basis of polymorphic fragments indicated a narrow variation in C. arietinum and recognized two major clusters in the genus Cicer. The first one included four species of sect. Monocicer: C. echinospermum, C. arietinum, C. reticulatum and Iranian material of C. bijugum. The second cluster contained annual and perennial species belonging to sections Chamaecicer, Polycicer and Acanthocicer. The character state of morphological and ecological traits on the RAPD phenogram indicated no monophyletic incision. Our results suggested that the high genetic difference between annual and perennial species might be regarded as a rapid speciation of section Monocicer. Reconsideration of traditional classification of sections Polycicer and Acanthocicer is necessary. The Desi and Kabuli types of C. arietinum could not be separately grouped at the DNA level, and the low level of genetic variation of C. arietinum may result from a bottleneck during the domestication process.     

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 diversity of Curcuma alismatifolia Gagnep.(Zingiberaceae) in Thailand as revealed by allozymepolymorphism

Allozyme polymorphism at seven loci (TPI, G6PD-2,IDH-1, SKD-2, MDH-1, GOT-1, andGOT-2) was employed to detect the level of geneticdiversity in C.alismatifolia populations from both cultivatedand wild habitats in Thailand. High diversity was observed in allpopulations with relatively lower values in cultivated populations.Percentage of polymorphic loci (P)varied from 85.7–100% in cultivated populations comparedwith 100% in all natural populations. Allele number per locus(A _L) was 3.14 in cultivatedpopulations, and from 2.86–4 in natural populations. Allelenumber per polymorphic locus(A _P) of cultivated andnatural populations ranged from 3.14–3.5 and 2.86–4,respectively. Genetic diversity within populations(H _S) varied from0.586–0.611 in cultivated and from 0.621–0.653 in naturalpopulations. The genetic identity(I _SP) for the species was0.833. The cultivated populations yielded higher value of geneticidentity with highland populations(I _{C /H} =0.776) than with the lowland ones(I _{C /L} =0.754). The analysis of genetic similarities with theNeighbor-Joining algorithm results in the separation ofcultivated populations from all wild populations. One highlandpopulation from the tourist spot, H2, was placed in a separatecluster between the cultivated and other wild populations. It isconsidered as the possible origin of the cultivatedpopulations.     

Genetic relationships among cultivated types of Perilla frutescens and their weedy types in East Asia revealed by AFLP markers

AFLP markers were employed to detect genetic diversity in two cultivated Perilla frutescens (i.e. var. frutescens and var. crispa) and their weedy types and to assess their genetic relationships. Analysis of 60 Perilla accessions from China, Korea and Japan by seven AFLP primer combinations identified a total of 125 fragments, of which 80 (64%) were polymorphic at the species level. The phenotypic diversity meassured by Shannon's index of information for the cultivated type of var. frutescens, the weedy type of var. frutescens, the cultivated type of var. crispa and the weedy type of var. crispa were on average 1.07, 2.23, 1.24 and 1.75, respectively. The weedy types exhibited high within-type variation in spite of a small number of samples. In the neighbor joining tree, two major clusters were recognized: (1) cultivated type of var. frutescens, (2) weedy type of var. frutescens, and cultivated and weedy types of var. crispa. The cultivated types of var. frutescens and of var. crispa were sharply separated by AFLPs. However, there remained ambiguities in regard to the intraspecific relaltionships, due to the clustering of the weedy types which occured in each of the clusters of the cultivated types. Two cultivated types of P. frutescens and their weedy types should be taxonomically considered as a P. frutescens complex. The present AFLP data are consistent with the hypothesis that China is the original place of cultivation of var. frutescens and Korea is a secondary center of diffusion of var. frutescens.     

Phylogenetic relationships among species of Citrullus and the placement of C. rehmii De Winter as determined by Internal Transcribed Spacer (ITS) sequence heterogeneity

The internal transcribed spacer regions (ITS1 and ITS2) of the 18S-25S nuclear ribosomal DNA from the four recognized species of Citrullus (C. lanatus var. lanatus (Thunb.) Matsum. & Nakai., C. lanatus var. citroides (Bailey) Mansf., C. colocynthis (L.) Schrad., C. ecirrhosus Cogn., and C. rehmii De Winter) and Acanthosicycos naudinianus (Sond.) C. Jeffrey were amplified by PCR, and direct sequenced. Within the taxa examined, the length of ITS1 varied from 216 bp to 219 bp, and ITS2 varied from 239 bp to 249 bp. The average %CG content ranged from 59 to 64% and from 62 to 66% for ITS1 and ITS2, respectively. The greater length variation observed in ITS2 was primarily attributable to the occurrence of a (CC)_n microsatellite. Cladistic (PAUP) and phenetic (MEGA) analyses resulted in highly resolutive trees. ITS sequence analysis placed the recently described C. rehmii adjacent to the cultivated watermelon and supported the validity of the species classification of this taxa.