Relationships Among <Emphasis Type="Italic">Brassica napus</Emphasis> (L.) Germplasm from Spain and Great Britain as Determined by RAPD Markers

The genetic diversity and the relationships among a collection of Brassica napus L. European populations were evaluated using random amplified polymorphic DNA markers. The study included 33 accessions of B. napus collected from Galicia (northwestern Spain) and 18 British cultivars, 16 accessions of B. napus and two accessions of Brassica oleracea L. used as controls. DNA from 25 individuals per population was analyzed using 18 decamer primers. One hundred thirty-eight amplification products were scored of which 105 were polymorphic. These bands ranged in size from 350 to 2500 base pairs. Similarity coefficients and cluster analysis were computed and six groups were obtained. Cluster I was the largest and included all the landraces from northwestern Spain, except two accessions that grouped separately into Clusters III and IV, respectively. A low level of genetic variability was detected among the B. napus Spanish genotypes, while considerable diversity was present among the British ones, which grouped into three groups, two main clusters and one group formed by one accession. Cluster II included all commercial varieties grown in Great Britain whereas Cluster V grouped local varieties maintained by the growers for many years. Cluster VI was a singularity formed by one entry. British accessions of B. oleracea had the greatest dissimilarity with all the other populations and grouped separately in Clusters VII and VIII. As conclusion, B. napus landraces used in northwestern Spain as leafy-green vegetable probably have an independent origin from B. napus crops grown in other European regions. Besides, separate domestication in northwestern Spain and Great Britain for a different end use might have led to two distinct gene pools.     

Genetic diversity in the Chamaecytisus proliferus complex (Fabaceae: Genisteae) in the Canary Islands in relation to in situ conservation

Summary Electrophoresis was carried out for six isozyme systems on 175 accessions of the seven morphological forms of Chamaecytisus proliferus (L. fil.) Link (Fabaceae: Genisteae) from the Canary Islands. Previous studies have shown that there is clear differentiation into morphological and ecological forms. The underlying genetic variation as visualised by studying isozymes does not reflect this. While substantial genetic diversity can be identified, this was found to be as much within as between morphological forms. This genetic diversity was found to decrease from east to west, and reflects the pattern of variation remaining after colonisation of the individual islands, which is assumed to have progressed in the same direction. Subsequently, adaptive radiation has given rise to the overlying morphological variation as exhibited in the seven morphological forms.In terms of in situ conservation, overall genetic diversity can be easily conserved in the abundant populations occurring in the east of the archipelago, while more attention is required for conservation of the much rarer morphological forms found in the west, despite their relative lack of isozyme diversity.Abbreviations ACO aconitase - ADH alcohol dehydrogenase - IDH Asocitrate dehydrogenase - MDH malate dehydrogenase - PGD 6-phosphogluconate dehydrogenase - PGM phosphoglucomutase - CA cluster analysis - PCA principal component analysis     

Characterisation and Variation of Morphological Traits and Storage Proteins in Spanish Emmer Wheat Germplasm (<Emphasis Type="Italic">Triticum Dicoccon</Emphasis>)

Emmer wheat is hulled wheat that was wide cultivated in Spain at the past. Actually, the most of this germplasm is conserved in Germplasm Banks, and only two small populations have been found in Asturias (North of Spain) in a recent collecting mission. In this work, a collection of 31 Spanish emmer lines developed from identical number of accessions of two Germplasm Banks was analysed for morphological spike traits and seed storage protein composition. Up to seven different botanical varieties were detected, which suggest the presence of a wide diversity, although lower than the historically described 10 botanical varieties. At level of seed storage proteins, the lines showed a high diversity, although the new alleles were present with low frequency in materials with scarce agronomic interest for the farmers (var. atratum, var. lagascae or var. pycnurum). This last circumstance could translate in a lost of variability by genetic drift.     

Genetic variation in wild and cultivated artichoke revealedby RAPD markers

Determination of intraspecific genetic diversity is an important first step for the utilization of genetic resources and canprovide useful information on crop evolution. A living collection of Italian and foreign artichoke varieties and ecotypes is maintained at the Germplasm Institute, Bari, Italy. A total of 32 accessions of cultivated (Cynara cardunculus L. var.scolymus (L.) Fiori), three of wild (Cynara cardunculus var.cardunculus L.) artichoke and two ofcultivated cardoon (Cynara cardunculus var.altilis L.) were analysed in order to studygenetic variation and relationships within the species, using RAPDmarkers. Cultivated accessions were selected according tomorphological variation and geographical distribution available inthe collection. Fifty arbitrary decamer primers were initially tested, 18 ofwhich showed from 3 to 10 unambiguously interpretable fragments. Anintra-accession analysis using 4 varieties and 8 polymorphicprimers revealed that no RAPD variation was detected amongindividuals. Jaccard's similarity index (JSI) was comprisedamong 1 and 0.693 when all accessions were considered, on the otherhand, within the cultivated artichoke, JSI ranged between 1 and0.817. A UPGMA dendrogram based on the similarity matrix showed thatwild artichokes were clearly separated from cultivated accessions.Moreover, within cultivated artichokes, several groups could bedistinguished.     

Genetic diversity in grasspea (Lathyrus sativus L.)

Genetic diversity was investigated in 348 accessions and subaccessions of grasspea (Lathyrus sativus L.) from 10 geographical regions. Polymorphism for 20 isozymes of 13 enzyme systems was studied to estimate the genetic diversity. The Near East and North Africa regions included the most variability for these isozyme systems, suggesting that the center of diversity (center of origin) for grasspea is in this general area. The lowest variability was found in accessions and subaccessions from South America, followed by those from Sudan–Ethiopia. Diversity was measured for individual loci over regions and EST-1 and SKDH had the highest genetic diversity. The closest genetic diversity was observed for LAP-2, followed by AAT-1 and PGM. The closest genetic distance existed between populations from the Near East and North Africa. Populations from South Asia and Sudan–Ethiopia, though geographically widely separated, exhibited a closer genetic distance from each other than from other regions.     

桂林地区蜈蚣蕨群体遗传多样性及与南宁地区群体的对比分析

采用等位酶电泳技术,研究了桂林地区的桂林市、雁山镇、永福县的野生蜈蚣蕨群体的遗传多样性,并与南宁地区群体进行比较。所测定的酶系统包括：氨基肽酶（AMP）、还原型辅酶Ⅰ心肌黄酶（DIA）、异柠檬酸脱氧酶（IDH）、苹果酸脱氨酶（MDH）、6-磷酸葡萄糖酸脱氢酶（PGD）、磷酸葡萄糖异构酶（PGI）、磷酸葡萄糖变位酶（PGM）和莽草酸脱氧酶（SKD共8个酶系统,15个酶位点。分析结果表明：桂林地区蜈蚣蕨群体内遗传多样性水平中等偏上,每个位点的等位基因平均为1．7,多态位点百分数为60％,平均期望杂合度为0．251,略高于南宁地区。     

Isozyme polymorphism provides fingerprints for germplasm ofArachis glabrata Bentham

Arachis glabrata Bentham, a wild, perennial relative of the cultivated peanut, has been under development as a forage plant for the subtropics and tropics since the early 1960s. To complement the limited genetic resources available in those years, a significant germplasm collection has been assembled in recent years. But little is known about the genetic diversity in this species because research has focused mainly on agronomy. This study aimed to investigate the utility of isozymes for characterisingA. glabrata germplasm. Polyacrylamide gel electrophoresis (PAGE) was applied to rhizome-tip tissue. Isozyme characterisation by PAGE showed a high degree of intraspecific polymorphism for the isozymes α-EST, ACP, GOT, and DIA. When analysed together, the four isozyme systems could clearly distinguish all 15 accessions ofA. glabrata held in the germplasm collection at CIAT. Although several accessions originated from the same region, similarity in isozyme patterns was not correlated with geographic origin. Examining rhizome-tip tissue is useful inA. glabrata, a species which produces few seeds.     

Assessment of genetic variation in a working collection of Vigna vexillata (L.) A. Rich. by isozyme and RAPD analyses

Genetic variation based on isozyme and RAPD analyses was investigated in 47 and 34 accessions respectively of Vigna vexillata from different geographical origins and belonging to three botanical varieties. A total of 9 enzyme systems were studied, accounting for 14 putative loci, 8 of which were polymorphic. The analysis of genetic diversity revealed a low level of within accession variation (H_S=0.013), while between accession diversity (D_ST) was 0.120. Coefficient of gene differentiation (G_ST) was 0.905, indicating that most variation was among accessions. Nei's genetic distances were calculated on the basis of allelic frequencies and a UPGMA dendrogram was constructed. Twenty arbitrary 10-mer oligonucleotides were used in RAPD analysis. Amplification profiles disclosed a higher level of polymorphism than isozymes. Based on amplification patterns, the similarity index of Jaccard was calculated and a dendrogram constructed on the basis of the similarity matrix. The final clustering based on RAPD data was similar to the one obtained using isozyme allelic frequencies. The classification in botanical varieties did not reflect the allelic constitution of the different samples. On the other hand, referring to geographical origin, most accessions from Africa and from Latin America were distributed respectively in two distinct clusters in the dendrogram. This grouping might also reflect the differences observed in the germination behaviour of V. vexillata from the two continents.     

Evaluation of a core collection of Brassica oleracea accessions for resistance to white rust of crucifers (Albugo candida) at the cotyledon stage

The identification of seedling resistance to white rust of crucifers was performed in a screening of a B. oleracea core collection with 400 accessions representing the genetic and geographic diversity of the species. Fifty seedlings per accession were tested against the Portuguese isolate Ac502 using the methodology and evaluation procedures developed by and . The percentage of resistant seedlings (%R) and the conventional rating criteria of the mean Disease Index (DI) based on the two different evaluation procedures of disease expression used, were compared and adopted as the criteria to rank the accessions for their interest as sources of resistance. A great variability of reactions was found between and within accessions of the core collection, ranging from complete resistance to full susceptibility. Sources of resistance were found namely among the cauliflowers, broccoli and tronchuda cabbages gene pools. Forty-seven accessions presented at least 20% of resistant seedlings. Nine accessions (the kales INRA18 and INRA62, the cauliflowers HRI4856, HRI4866 and HRI5424, the loose-head cabbage HRI11555, the savoy cabbage BRA848, the black broccoli HRI6318 and the Portuguese tronchuda cabbage ISA207) presented 50–78% of resistant seedlings and so they should be considered as potential and useful sources for direct use in breeding programs for white rust resistance. Fourteen inbred lines, representing the full range of disease expression, derived from resistant accessions of the core collection were also tested for resistance to other two Portuguese isolates (Ac503 and Ac504) and to a UK isolate. The results provided no evidence of differential reaction to the A. candida isolates tested.     

Analysis of Genetic Diversity in the Brassica napus L. Gene Pool Using SSR Markers

Genetic diversity throughout the rapeseed (Brassica napus ssp. napus) primary gene pool was examined by obtaining detailed molecular genetic information at simple sequence repeat (SSR) loci for a broad range of winter and spring oilseed, fodder and leaf rape gene bank accessions. The plant material investigated was selected from a preliminary B. napus core collection developed from European gene bank material, and was intended to cover as broadly as possible the diversity present in the species, excluding swedes (B. napus ssp. napobrassica (L.) Hanelt). A set of 96 genotypes was characterised using publicly available mapped SSR markers spread over the B. napus genome. Allelic information from 30 SSR primer combinations amplifying 220 alleles at 51 polymorphic loci provided unique genetic fingerprints for all genotypes. UPGMA clustering enabled identification of four general groups with increasing genetic diversity as follows (1) spring oilseed and fodder; (2) winter oilseed; (3) winter fodder; (4) vegetable genotypes. The most extreme allelic variation was observed in a spring kale from the United Kingdom and a Japanese spring vegetable genotype, and two winter rape accessions from Korea and Japan, respectively. Unexpectedly the next most distinct genotypes were two old winter oilseed varieties from Germany and Ukraine, respectively. A number of other accessions were also found to be genetically distinct from the other material of the same type. The molecular genetic information gained enables the identification of untapped genetic variability for rapeseed breeding and is potentially interesting with respect to increasing heterosis in oilseed rape hybrids.     

Genetic characterization of a collection of chayote, <Emphasis Type="Italic">Sechium edule</Emphasis> (Jacq.) Swartz,in Costa Rica by using isozyme markers

We established protocols for the analysis of genetic diversity in chayote (Sechium edule) by using isozyme markers, thereby determining the level of genetic diversity present in 42 accessions of chayote from Costa Rica. We obtained clear and reproducible zymograms for eight enzyme staining systems: PGM, 6-PGD, PGI, IDH, MDH, SOD, SKD, and EST, and were able to score 14 putative loci. Eight of the 14 loci examined were polymorphic. We found 35 distinct multilocus genotypes among these accessions. Five of these multilocus genotypes were homozygous for all loci. In addition, our data also revealed that most of the multilocus genotypes (24) were heterozygous for only one of the eight loci, and the rest were heterozygous for two or three loci (9 and 4 accessions, respectively). Seven multilocus genotypes were found in two different accessions. Dice similarity coefficient was used to study the relationship between accessions. This analysis, based on the presence and absence of alleles, revealed that accessions collected in the same location seldom shared the same multilocus genotype. The value of isozyme polymorphisms as tools to continue studies on the characterization of chayote is discussed.     

Strategies and priorities in field collections for ex situ conservation: the case of the Israel Plant Gene Bank

The ex situ collection of the Israel Plant Gene Bank (IGB) aims to encompass the rich local flora and its genetic diversity with an emphasis on crop wild relatives. However, to properly establish a core collection, collection efforts must be prioritized and strategized. We previously classified local plant genetic resources into four priority groups that assisted in strategizing the collection activities. The following years of intensive collection activity yielded over 4200 banked accessions. However, these do not necessarily represent the distribution range of the target species for collection (TSC) and consequently, their genetic diversity. To best cover the latter, the collecting area was divided into botanical districts and the magnitude of the collection was determined according to prioritization group, e.g., a wild relative of an agricultural crop with a vast distribution range should be represented by a larger number of banked accessions than one with a smaller range. Continuous evaluation of specific needs shapes the collection scheme of the IGB to maximize collection efforts, better represent the presumed genetic diversity of TSC, and establish its core collection.     

A wide range of genetic diversity in pear (<Emphasis Type="Italic">Pyrus ussuriensis</Emphasis> var. <Emphasis Type="Italic">aromatica</Emphasis>) genetic resources from Iwate,Japan revealed by SSR and chloroplast DNA markers

Iwateyamanashi (Pyrus ussuriensis var. aromatica) is one of the Pyrus species which grows wild in Japan. The number of Iwateyamanashi trees has been decreasing, so conservation and evaluation is urgently needed. Over 500 accessions of Pyrus species collected from Iwate in northern Tohoku region are maintained at Kobe University as an Iwateyamanashi germplasm collection. In order to investigate the genetic diversity, five SSR (simple sequence repeat) markers, developed from Japanese and European pear were examined for 86 Pyrus individuals including 58 accessions from Iwate. These SSR loci could discriminate between all the Iwate accessions except for 10 that bear seedless fruit, as well as determine the genetic diversity in Iwateyamanashi germplasms. High levels of variation were detected in 41 alleles and the mean observed heterozygosity across 5 loci was 0.50 for the Iwate accessions. Seedless accessions sharing identical SSR genotype with the local pear variety “Iwatetanenashi” were supposed to have been propagated vegetatively via grafting. In an UPGMA phenogram, Japanese pear varieties (P. pyrifolia) were clustered into two groups with some Iwate accessions including seedless ones. Another 38 Iwate accessions were not clustered clearly, and there was no clear relationship between these accessions and geographical distribution or morphological characters. Allele frequency revealed that the Iwate accessions were genetically more divergent than the Japanese pear varieties. Most Japanese pears possessed a 219 bp deletion at a spacer region between the accD and psaI genes in the chloroplast DNA (cpDNA), but other Pyrus species and two Iwateyamanashi trees did not. In the Iwate accessions, 79.3% had a deletion type cpDNA and others had a standard type cpDNA without deletion. These results are indicative of the wide range of genetic diversity in the Iwate accessions which include Japanese pear varieties. A combination of SSR and cpDNA analyses revealed high heterogeneity in Iwateyamanashi and coexistence of Iwateyamanashi and hybrid progeny with P. pyrifolia. These could be reasons for the wide range of continuous morphological variation described previously.     

Analysis of genetic diversity in a sweet potato (Ipomoea batatas L.) germplasm collection from Tanzania as revealed by AFLP

Sweet potato (Ipomoea batatas L.) is the fifth most important crop in the developing countries after rice, wheat, maize and cassava. The amplified fragment length polymorphism (AFLP) method was used to study the genetic diversity and relationships of sweet potato accessions in the germplasm collection of Sokoine University of Agriculture, Morogoro and Sugarcane Research Institute, Kibaha, Tanzania. AFLP analysis of 97 sweet potato accessions using ten primer combinations gave a total of 202 clear polymorphic bands. Each one of the 97 sweet potato accessions could be distinguished based on these primer combinations. Estimates of genetic similarities were obtained by the Dice coefficient, and a final dendrogram was constructed with the un-weight pair-group method using arithmetic average. AFLP-based genetic similarity varied from 0.388 to 0.941, with a mean of 0.709. Cluster analysis using genetic similarity divided the accessions into two main groups suggesting that there are genetic relationships among the accessions. Principal Coordinate analysis confirmed the pattern of the cluster analysis. Analysis of molecular variance revealed greater variation within regions (96.19%) than among regions (3.81%). The results from the AFLP analysis revealed a relatively low genetic diversity among the germplasm accessions and the genetic distances between regions were low. A maximally diverse subset of 13 accessions capturing 97% of the molecular markers diversity was identified. We were able to detect duplicates accessions in the germplasm collection using the highly polymorphic markers obtained by AFLP, which were found to be an efficient tool to characterize the genetic diversity and relationships of sweet potato accessions in the germplasm collection in Tanzania.     

Perennial kales: collection rationalization and genetic relatedness to other Brassica oleracea crop types

Perennial kale is a rare leafy vegetable and forage crop that is mainly vegetatively propagated and therefore expensive to conserve ex situ. A genebank collection of 47 perennial kales and 34 reference samples from the main Brassica oleracea crop types were characterized with seven microsatellite markers in order to verify potential redundancies and to obtain more insight in the position of perennial kales within B. oleracea. Based on the obtained results and on data from previous studies, the collection was reduced with 49% to 24 perennial kales. Considering this level of reduction, it was estimated that the investments made for the final verification by microsatellite analysis are returned after only 4-year time. A principal coordinate plot clearly separated the perennial kales from the other crop types of B. oleracea, except in one case. This deviating accession of vegetatively preserved perennial kale clustered closely together with the single seed-preserved accession of perennial kale included in the study. These two accessions occupied an intermediate position between the group of vegetatively propagated perennial kales and the group of seed-propagated Brassica accessions, suggesting a hybridization background with another B. oleracea crop type. The microsatellite study demonstrated a close genetic relationship among the investigated perennial kales and their unique position within B. oleracea.     

Genetic relationships of Portuguese coles and other close relatedBrassica genotypes using nuclear RFLPs

Nuclear RFLPs were used to study the genetic relationships of 2 Portuguese coles, tronchuda cabbage and Galega kale, and 13 otherBrassica oleracea cultivars and 4 nine-chromosome wild brassicas. Cluster and principal coordinates analysis were conducted using RFLP data from 60 probe-enzyme combinations, detecting 277 polymorphic restriction fragments. The results showed that the accessions clustered in five groups: one with all theB. oleracea cultivars except kailan, and the four others isolated with kailan, wildB. oleracea, B. insularis andB. cretica, andB. montana, respectively. Kailan was separated from the other accessions ofB. oleracea cultivars and genetically close to the wildB. oleracea, that was clearly separated from the other nine-chromosome wild brassicas. In theB. oleracea cultivars 3 groupings were clearly individualized: i) including broccolis and cauliflower; ii) with a misture of kales and cabbages originally from Central-North Europe; iii) formed by Portuguese coles. These preliminary results suggest the existence of three major regions of domestication of B. oleracea in Europe: Italy, Central-North Europe and Portugal. Kailan or chinese kale seems to have evolved separately from the otherB. oleracea cultivars in Eastern Asia.     

Database derived microsatellite markers (SSRs) for cultivar differentiation in <Emphasis Type="Italic">Brassica oleracea</Emphasis>

Fifty-nine Brassica oleracea cultivars, belonging to five botanical varieties, were evaluated for microsatellite (SSR) polymorphisms using 11 database sequence derived primer pairs. The cultivars represented 12 broccoli (Brassica oleracea var. italica), ten Brussels sprouts (B. o. var. gemmifera), 21 cabbage (B. o. var. capitata, including the groups white and red cabbage), six savoy cabbage (B. o. var. sabauda), and ten cauliflower (B. o. var. botrytis) cultivars from 13 seed suppliers. The 11 primer pairs amplified in total 47 fragments, and differentiated 51 of the cultivars, whereas the remaining eight cultivars were differentiated from the rest in four inseparable pairs. All SSR markers, except one, produced a polymorphic information content (PIC value) of 0.5 or above. The average diversity for all markers within the tested material was 0.64. There was no major difference in the diversity within botanical varieties and groups. The cluster analysis and the resulting dendrogram showed that the cultivars tended to group within these taxonomic units. The present study substantiates the use of microsatellite markers as a powerful tool for cultivar differentiation and identification in vegetable brassicas.     

Identification and genetic relationships of kenaf (Hibiscus cannabinus L.) germplasm revealed by AFLP analysis

Kenaf (Hibiscus cannabinus L.) is one of the world's most economically important fiber crops. In order to identify different varieties, and investigate its diversity and genetic relationships, twenty-three kenaf accessions and two accessions of its relative, roselle (H. sabdariffa var. altissima), were analyzed by morphological characterization and AFLP fingerprinting. It is very difficult to identify kenaf accessions based merely on morphological characters, due to their limited variation. For the AFLP study, a total of 505 polymorphic markers (out of 560) were produced by six selected AFLP primer combinations. The AFLP fingerprinting was effective in identifying all kenaf accessions included in the study. Kenaf and roselle are independent species with close relationships, and great genetic diversity was also detected among the kenaf accessions with different origins, based on the analysis of the AFLP markers. The AFLP analysis strongly supports the opinion that kenaf originated in Africa. It also demonstrated that the dissemination of kenaf was from Africa through Asia to Central and North America.