Genetic diversity of Swiss maize (<Emphasis Type="Italic">Zea mays</Emphasis> L. ssp. <Emphasis Type="Italic">mays</Emphasis>) assessed with individuals and bulks on agarose gels

About 65 years ago, more than 150 Swiss maize landraces (Zea mays L. ssp. mays) of the flint type were collected and conserved ex situ. Due to the climatically and culturally diverse environment of the Alps, a considerable genetic diversity of this material was assumed. To prove this, an efficient method was required to carry out genetic profiling of all the accessions in the Swiss Gene Bank. Simple sequence repeat marker (SSR) profiling in combination with the visualization of the polymerase chain reaction (PCR) products on agarose gels was chosen. Here a set of 19 different landrace accessions was analyzed to: (i) investigate their genetic diversity, (ii) investigate and display the population structure and (iii) determine whether DNA bulks rather than single plants can be used for such analyses. Four repeated samples of one accession were found to be much closer to one another than to the rest of accessions. Furthermore, specific alleles were identified for several accessions. The PCR products of the bulked DNA samples represented only a small part of the variation revealed by the analysis of individuals. Loci with four base repeat motifs performed better in the analysis of bulks than loci with other repeat motifs. The correlation between genetic distance matrices, based on the analysis of individuals and bulks, respectively, was significant. Thus, the single plant approach allowed for sufficient differentiation of accessions, and DNA bulks visualized on agarose gels led to correlated genetic distances although a limited number of alleles were detected. Although the limited resolution of agarose gels likely causes some bias, profiling of larger sets with the individual plant approach appears feasible and more informative compared to the bulk analysis we conducted.     

Maize diversity in southern Brazil: indication of a microcenter of <Emphasis Type="Italic">Zea mays</Emphasis> L.

‘Microcenters of diversity’ are restricted geographical areas within which a significant diversity of genetic resources is accumulated. Maize genetic variability, one of the largest among cultivated species, particularly in far western Santa Catarina, southern Brazil, has shown a rich store of landraces, conserved on farm by small-scale farmers. Thus, this study aimed to characterize the diversity of Zea mays L. landraces in two municipalities of this micro-region in Santa Catarina, as well as its geographic distribution, developing the census of diversity as a methodological proposal. Diagnosis was made according to the farmers’ knowledge. The field survey was conducted in 70 rural communities involving 2049 farms. The methodology allowed identifying 136 populations of wild relatives, of which some belong to the species Zea luxurians (Durieu and Ascherson) Bird, and 1513 populations of maize landraces, comprising 1078 of popcorn, 337 of common maize, 61 of sweet maize and 37 of flour maize. The identification of 59 morphological groups, the high use, adaptive and agronomic values, and the Shannon index (H′), estimated according to the grain shape (0.79), endosperm type (0.73), size (0.87), color (1.40), and morphological group (3.16), showed at set a considerable diversity distribution conserved on farm in this micro-region. Landrace richness and the presence of wild relative species, associated with the local human activity and sociocultural aspects, allowed to characterize the far western Santa Catarina as a ‘microcenter of diversity’ of Zea mays L.     

AFLP analysis of genetic diversity in five accessions of Polish runner bean (<Emphasis Type="Italic">Phaseolus coccineus</Emphasis> L.)

Runner bean (Phaseolus coccineus L.) is traditionally cultivated in Poland for dry seeds. The national collection of runner bean maintained in the National Center for Plant Genetic Resources gathers 152 accessions, which are mainly landraces originated in Poland (68%), Ukraine (17%) and Slovakia (10%). The collection contains valuable genetic resources for bean breeders and research. The aim of this study was to describe the level and structure of genetic diversity of three landraces and two commercial cultivars of runner bean from the national collection in order to assess their genetic potential for breeding. Amplified Fragment Length Polymorphism analysis included five combinations of selective primers. Analysis of seven genetic diversity parameters reveled fair amount of genetic variation both in landraces and cultivars. High genetic diversity of commercial cultivars relative those of landraces suggests that the breeding process leading to their release was rather moderate and most likely included domestic gene pool of runner bean. Low gene diversity and low Nei’s genetic distance values as well as intergradations among accessions in the PCoA may indicate reduced variability P. coccineus grown in Poland as a result of its migration pathways.     

AFLP analysis of genetic diversity in leafy kale (<Emphasis Type="Italic">Brassica oleracea</Emphasis> L. convar. <Emphasis Type="Italic">acephala</Emphasis> (DC.) Alef.) landraces,cultivars and wild populations in Europe

AFLP markers were used to characterize diversity and asses the genetic structure among 17 accessions of kale landraces, cultivars and wild populations from Europe. The range of average gene diversity in accessions was 0.11–0.27. Several landraces showed higher levels of diversity than the wild populations and one cultivar had the lowest diversity measures. The landraces that were most genetically diverse were from areas where kales are known to be extensively grown, suggesting in situ conservation in these areas as a supplement to storage of seeds in gene banks. An analysis of molecular variance (AMOVA) showed that 62% of the total variation was found within accessions. For most accessions, genetic distance was not related to geographic distance. Similarities among accessions were probably not caused by recent gene flow since they were widely separated geographically; more likely the relationship among them is due to seed dispersal through human interactions. Our results indicate that a kale population found in a natural habitat in Denmark was probably not truly wild but most likely an escape from a cultivated Danish kale that had subsequently become naturalized.     

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.     

Comparison of the genetic diversity between <Emphasis Type="Italic">Triticum aestivum</Emphasis> ssp. <Emphasis Type="Italic">tibetanum</Emphasis> Shao and Tibetan wheat landraces (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) by using intron-splice junction primers

In order to evaluate and compare the germplasm resources of wheat in Tibet, we analyzed the genetic diversity of 136 Triticum aestivum ssp. tibetanum Shao and 119 Tibetan wheat landraces (Triticum aestivum L.) by using Intron-Splice Junction (ISJ) primers. The results showed that polymorphism of PCR products were obtained by 33 primer combinations, which accounted for 11% of the 300 primer combinations produced by 26 ISJ primers. A total of 333 stable bands can be amplified from the T. aestivum ssp. tibetanum Shao and 243 bands were polymorphic, which accounted for 72.9% of the total bands. Tibetan wheat Landraces produced 316 stable bands, of which 197 bands were polymorphic. The polymorphic bands accounted for 62.34% of the total bands produced from Tibetan wheat landraces. The genetic diversity of T. aestivum ssp. tibetanum Shao was higher than that of Tibetan wheat landraces in Tibet, suggesting that T. aestivum ssp. tibetanum Shao can be used as important genetic resource for the breeding and genetic improvement of wheat in Tibet. Matrix (1, 0) was generated according to the presence or absence of the bands produced from a particular wheat accession. Clustering and principle coordinates analysis showed that T. aestivum ssp. tibetanum Shao and Tibetan wheat landraces were divided into two groups. We conclude that high polymorphisms produced by ISJ primers can reflect the genetic diversity between T. aestivum ssp. tibetanum Shao and Tibetan wheat landraces.     

Morphological characterization of snake melon (<Emphasis Type="Italic">Cucumis melo</Emphasis> var. <Emphasis Type="Italic">flexuosus</Emphasis>) populations from Palestine

Genetic diversity in 50 snake melon accessions collected from Palestine (West Bank) was assessed by examining variation in 17 phenotypic characters. These accessions belonged to four important landraces of Cucumis melo var. flexuosus: Green “Baladi” (GB), white Baladi (WB), green Sahouri (GS), and white Sahouri (WS). Principal component analysis (PCA) and a dendrogram were performed to determine relationships among populations and to obtain information on the usefulness of those characters for the definition of cultivars. PCA revealed that secondary fruit skin color, flesh color, primary fruit skin color, and secondary skin color pattern were the principal characters to discriminate melon accessions examined in the present study. According to the scatter diagram and dendrogram, landraces of C. melo var. flexuosus: GB, WB, GS, and WS formed different clusters. However, based on Euclidean genetic coefficient distances, GB and WB had the least degree of relatedness with GS and WS, indicating distantly related landraces (Baladi and Sahouri). On the other hand, the highest degree of relatedness was detected between WS on one hand, and both GS and WB on the other indicating closely related cultivars. Fruit traits variability among the different snake melon landraces was evaluated and discussed in this study. This evaluation of fruit trait variability can assist geneticists and breeders to identify populations with desirable characteristics for inclusion in cultivars breeding programs.     

Detection of genetic integrity of conserved maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.) germplasm in genebanks using SNP markers

Twenty maize landrace accessions regenerated and conserved in five maize genebanks were investigated for genetic integrity using 1,150 Single Nucleotide Polymorphisms (SNPs) and 235 SNP haplotypes. The genetic diversity of three accessions changed significantly in terms of the average number of alleles per locus. Ten out of twenty accessions had significantly different SNP allelic frequencies, either after regeneration or in the same accession held in different genebanks. The proportion of loci with significant changes in SNP allelic frequency was very low (37/1,150). Changes in the major allelic frequency (MAF) for the majority of SNP loci (60.2–75.2%) were less than 0.05. For SNP haplotypes, the genetic diversity of four accessions changed significantly in terms of average number of haplotype alleles and polymorphic information content (PIC) per locus. The proportion of SNP haplotype alleles lost in the later generations ranged between 0 and 22.6%, and at the same time 0–19.9% of the SNP haplotype alleles appeared in later generations, however, these were absent in the earlier generations. Dynamic changes in genetic integrity, in terms of presence and absence of genes (alleles), by both SNP and SNP haplotype analysis were detected during regeneration. A suboptimum number of ears harvested in one generation can be combined with those from another, repeated regeneration to capture the diversity of the previous generation. Use of molecular markers during regeneration of accessions can help in understanding the extent of genetic integrity of the maize accessions in ex situ genebanks and in recommending the best practice for maintaining the original genetic diversity of the genebank accessions.     

Variation in melon (<Emphasis Type="Italic">Cucumis melo</Emphasis>) landraces adapted to the humid tropics of southern India

We present here the first comprehensive genetic characterization of melon landraces from the humid tropics of southern India. The genetic diversity among 50 melon landraces collected from 3 agro-ecological regions of southern India (6 agro-ecological sub-regions) was assessed by variation at 17 SSR loci, morphological traits of plant habit and fruit, 2 yield-associated traits, pest and disease resistance, biochemical composition (ascorbic acid, carotenoids, titrable acidity) and mineral content (P, K, Fe, Zn). Differences among accessions were observed in plant and fruit traits. Melon germplasm with high titrable acidity, higher than average amounts of mineral content and resistance to Cucumber mosaic virus, Zucchini yellow mosaic virus, powdery mildew (races 1, 2, 3, 5), Fusarium wilt (races 1, 2), Aphis gossypii and leafminer was recorded in the collection. A high level of genetic variability in melon germplasm was suggested by the SSR analysis. Comparative analysis using SSRs of the genetic variability between Indian melons from north, south, and east regions and reference accessions of melon from Spain, France, Japan, Korea, Iraq, Zambia showed regional differentiation between Indian melon accessions and that Indian germplasm was weakly related to the melon accessions from other parts of the world, suggesting that an important portion of the genetic variability found within this melon collection has not been used yet for the development of new cultivars. Additional collections of acidulus melon germplasm should be made in southern India and adequate management of this important genetic resource is clearly a necessity.     

The genetic characterization of Turkish watermelon (<Emphasis Type="Italic">Citrullus lanatus</Emphasis>) accessions using RAPD markers

Genetic diversity of the Turkish watermelon genetic resources was evaluated using different Citrullus species, wild relatives, foreign landraces, open pollinated (OP) and commercial hybrid cultivars by RAPD markers. The germplasm was consisted of 303 accessions collected from various geographical regions. Twenty-two of 35 RAPD primers generated a total of 241 reproducible bands, 146 (60.6%) of which were polymorphic. Based on the RAPD data the genetic similarity coefficients were calculated and the dendrogram was constructed using UPGMA (Unweighted pair-group method with arithmetic average). Cluster analysis of the 303 accessions employing RAPD data resulted in a multi-branched dendrogram indicating that most of the Turkish accessions belonging to var. lanatus of Citrullus lanatus (Thunb.) Matsum et Nakai were grouped together. Accessions of different Citrullus species and Praecitrullus fistulosus (Stocks) Pangalo formed distant clusters from C. lanatus var. lanatus. Among 303 accessions, a subset of 56 accessions was selected representing different groups and a second dendrogram was constructed. The genetic similarity coefficients (GS) within the Turkish accessions were ranged from 0.76 to 1.00 with 0.94 average indicating that they are closely related. Taken together, our results indicated that low genetic variability exist among the watermelon genetic resources collected from Turkey contrary to their remarkable phenotypic diversity.     

Exploitation of <Emphasis Type="Italic">Malus</Emphasis> EST-SSRs and the utility in evaluation of genetic diversity in <Emphasis Type="Italic">Malus</Emphasis> and <Emphasis Type="Italic">Pyrus</Emphasis>

A total of 8117 suitable SSR-contaning ESTs were acquired by screening from a Malus EST database, among which dinudeotide SSRs were the most abundant repeat motif, within which, CT/TC followed by AG/GA were predominant. Based on the suitable sequences, we developed 147 SSR primer pairs, of which 94 pairs gave amplifications within the expected size range while 65 pairs were found to be polymorphic after a preliminary test. Eighteen primer pairs selected randomly were further used to assess genetic relationship among 20 Malus species or cultivars. As a result, these primers displayed high level of polymorphism with a mean of 6.94 alleles per locus and UPGMA cluster analysis grouped twenty Malus accessions into five groups at the similarity level of 0.6800 that were largely congruent to the traditional taxonomy. Subsequently, all of the 94 primer pairs were tested on four accessions of Pyrus to evaluate the transferability of the markers, and 40 of 72 functional SSRs produced polymorphic amplicons from which 8 SSR loci selected randomly were employed to analyze genetic diversity and relationship among a collection of Pyrus. The 8 primer pairs produced expected bands with the similar size in apples with an average of 7.375 alleles per locus. The observed heterozygosity of different loci ranged from 0.29 (MES₉₆) to 0.83 (MES₁₃₈), with a mean of 0.55 which is lower than 0.63 reported in genome-derived SSR marker analysis in Pyrus. The UPGMA dendrogram was similar to the previous results obtained by using RAPD and AFLP markers. Our results showed that these EST-SSR markers displayed reliable amplification and considerable polymorphism in both Malus and Pyrus, and will contribute to the knowledge of genetic study of Malus and genetically closed genera.     

Salt tolerance in <Emphasis Type="Italic">Zea mays</Emphasis> (L). following inoculation with <Emphasis Type="Italic">Rhizobium</Emphasis> and <Emphasis Type="Italic">Pseudomonas</Emphasis>

This study aimed to investigate the effect of inoculation with plant growth-promoting Rhizobium and Pseudomonas species on NaCl-affected maize. Two cultivars of maize (cv. Agaiti 2002 and cv. Av 4001) selected on the basis of their yield potential were grown in pots outdoors under natural conditions during July. Microorganisms were applied at seedling stage and salt stress was induced 21 days after sowing and maintained up to 50% flowering after 120 days of stress. The salt treatment caused a detrimental effect on growth and development of plants. Co-inoculation resulted in some positive adaptative responses of maize plants under salinity. The salt tolerance from inoculation was generally mediated by decreases in electrolyte leakage and in osmotic potential, an increase in osmoregulant (proline) production, maintenance of relative water content of leaves, and selective uptake of K ions. Generally, the microbial strain acted synergistically. However, under unstressed conditions, Rhizobium was more effective than Pseudomonas but under salt stress the favorable effect was observed even if some exceptions were also observed. The maize cv. Agaiti 2002 appeared to be more responsive to inoculation and was relatively less tolerant to salt compared to that of cv. Av 4001.     

Realizing value of genetic resources of <Emphasis Type="Italic">Allium</Emphasis> in India

The genetic resources of Allium in India are potential source of genes for widening the crop genetic base. Despite their high economic value, limited number of germplasm accessions of wild species have been collected and conserved mainly due to difficult access to areas of occurrence. The present work mainly highlights information on species diversity distribution and utilization of wild Allium species to assess the value of Indian germplasm and prioritization for future collection and conservation programmes.     

Maintenance of Squash (<Emphasis Type="Italic">Cucurbita</Emphasis> spp.) Landrace Diversity by Farmers' Activities in Mexico

Squash (Cucurbita spp.) is a common component in traditional cropping systems in Mexico, mainly in the agroecosystem known as the “milpa”, in which squash is cultivated in association with maize (Zea mays), the main crop. Using a questionnaire, 80 farmers were interviewed about crop production and selection practices in order to understand how these factors affect genetic diversity of local squash populations. We found that the most of the farmers who cultivate squash were elderly 59.8 ± 14.5 (mean ± SD; n = 78) years old. Squash varieties in the area were exclusively locally adapted landraces, and had not been replaced by modern squash cultivars. Two cultivated squash species, C. argyrosperma ssp. argyrosperma and C. moschata, had been grown intercropped with maize by 97.5% of the interviewed farmers, but only 50.0% were still producing squash at the time of the study. Farmers recognize typical characteristics of particular varieties within each of the local cultivated squash species, and selection is directed to maintain their identity. Nearly two thirds of the farmers (62.0%) had exchanged seeds of squash for planting, a practice that serves to increase genetic variability in the populations. All of the interviewed farmers were conscious of the possible hybridization between the wild gourd (C. argyrosperma ssp. sororia) and their cultivated squash. Despite various natural and human managed factors identified as contributing to enhancement of genetic diversity in these populations, results of the study show that genetic erosion of Cucurbita is likely in the region in the near future.     

Genetic diversity in oil and vegetable mustard (<Emphasis Type="Italic">Brassica</Emphasis><Emphasis Type="Italic">juncea</Emphasis>) landraces revealed by SRAP markers

Mustard (Brassica juncea) is an important crop in both ancient and modern world. It has a broad resource of genetic diversity that is used primarily as oilseed but as vegetables, condiment and medicines also. Its superior tolerance to adverse environments, e.g., drought, high temperature and low fertility suggests its better adaptability in future possible harsh environments. Chinese vegetable mustard displays a wide spectrum of morphotypes. A collection of 95 accessions of B. juncea representing oil and vegetable mustards from China, France, India, Pakistan, and Japan were assessed to determine diversity at the molecular level using sequence-related amplified polymorphism (SRAP). Eight SRAP primer combinations identified a total of 326 scorable fragments of which 161 were polymorphic (49.39%). The percentage of polymorphism for each primer combination varied from 21.88 to 66.67%. Both Shannon-Weaver and Simpson genetic diversity index indicated that the level of genetic diversity within vegetable mustard is much higher than within oil mustard, and also winter oil mustards are genetically more diverse than spring oil mustards. Based on the Cluster and Principal Coordinates analysis, which were conducted on the similarity matrix of SRAP marker data, vegetable, spring oil and winter oil mustard were clearly divided into three distinct groups and among these three groups, spring and winter oil mustard are geneticlly closer than vegetable mustard. This suggests that bilateral gene exchange between oil and vegetable gene pools in the breeding program will effectively elevate the genetic potential in developing higher yields, more disease resistance, better quality and better adapted lines.