Genetic variation and species relationships in Himalayan buckwheats as revealed by SDS PAGE of endosperm proteins extracted from single seeds and RAPD based DNA fingerprints

SDS PAGE of endosperm proteins and␣RAPD profiles from different accessions of Himalayan buckwheat were studied to determine their genetic variation and phylogenetic relationships. Comparisons based on Jaccard’s coefficient and UPGMA clustering revealed interrelationships broadly in conformity with conventional treatments. Cluster analysis of the endosperm protein profiles of the selected accessions revealed three broad clusters. A moderate level of intraspecific variability was detected in the endosperm protein profile of different accessions of Fagopyrum esculentum, Moench. Three subgroups were detected in cluster 1. Subgroup 1 included varieties designated as Local, Kamroo local, OC-2 and␣VL-7 which were collected from VPKAS, Almora. Local, Kamroo local and OC-2 showed a similarity coefficient of 1.0 inspite of their being identified as different accessions. VL-7 emerged out separately from the rest of the three accessions. Accessions having winged grains and those having striations on the seed coat formed a 2nd and 3rd subgroup, respectively. IC-13145 which been identified as “F. himalianum”, showed 100%␣similarity in endosperm protein profile with IC-13376 (F. esculentum) and 85–90% similarity which other accessions of F. esculentum. Our results indicate that “F. himalianum” belongs to the esculentum group and should not be regarded as a different species. Cluster 2 included all the accessions of F. tataricum (L.) Gaertn. with Sangla 1, 2, 3, 5, 6, 7 and IC-412863 showing 100% similarity. F. cymosum emerged as a separate group distinct from both esculentum and tataricum. Accessions of F. tataricum and F. cymosum did not show significant intraspecific variation in the SDS PAGE profile of endosperm proteins. Out of the 20 primers used, 3 generated robust, easily interpretable amplified products. While a 1490 bp and a 300 bp RAPD was detected only in F. tataricum, a 1154 bp RAPD was detected in all accessions of F. tataricum except in Shimla B-1. This variety is early maturing and has high seed yields.     

Diversity characterisation of broccoli (Brassica oleracea L. var. italica Plenck) landraces for their on-farm (in situ) safeguard and use in breeding programs

Brassica oleracea L. is an important species that originated in the eastern Mediterranean area. From there, B. oleracea L. subsp. capitata (L.) DC. convar. botrytis (L.) Alef. var. italica Plenck (broccoli) was introduced into Italy where considerable diversification took place. Several landraces of broccoli, which are highly appreciated by local people for their organoleptic traits, are presently cultivated in both large fields and home gardens. In this study, 10 landraces, 4 synthetics derived from landraces and 4 of the most used F1 hybrids were characterised for morpho-physiological and molecular traits with the principal aim of feeding information into the landrace protection schemes being developed in Italy and into breeding programs. The landraces and derived synthetics are well differentiated from each other and from the F1 hybrids for important agronomic and genetic traits. The principal component analysis based on analysis of morpho-physiological traits showed that overall the 2 main components accounted for 67.74 % of the total variation. The neighbour joining tree, based on 23 microsatellite markers, grouped accessions into 2 main clusters. One includes one synthetic variety and 2 F1 hybrids, while the other cluster includes all the other accessions. There were also several sub clusters. Most of the materials examined showed within-accession diversity. The information gathered in this study can be used to protect landraces through the schemes presently being developed in Italy and to register them in the European common catalogue of “conservation varieties”. In situ (on-farm) conservation of landrace diversity is discussed with reference to landrace use.     

Analyses of the seed protein contents on the cultivated and wild buckwheat <Emphasis Type="Italic">Fagopyrum esculentum</Emphasis> resources

The seed protein contents of 179 accessions belonging to ten species of genus Fagopyrum were determined by means of the Kjeldahl method. The results indicated that there are significant differences of seed protein content among different accessions within the same species. The average protein content of F. esculentum is 12.94%, ranging from 8.81–18.71%, and the protein content of Sibano is the highest, up to 18.71%, Gantian 1 the lowest with 8.81%. The average protein content of tartary buckwheat is 12.17%, with the range from 7.82–18.94%, and the protein content of cultivated tartary buckwheat Qianku 2 is the highest (18.59%), and that of Ganku 1 (7.82%) the lowest. Moreover, there are significant differences of seed protein content among different buckwheat species and between the different buckwheat types (between common buckwheat and tartary buckwheat, between the diploid and the tetraploid, between the cultivated and the wild, and between the big-achene group and the small-achene group). The average seed protein content of F. giganteum is the highest in all tested species, up to 17.81% with the range of 13.91–21.27%, and that of F. callianthum the lowest, only 8.31%. This study was financed by the Natural Science Foundation of China (30270852, 30471116), Program for New Century Excellent Talents in University (NCET-2004-0913), Guizhou Key International Cooperation Project (Qianke Hewai G Zi 2005#400108), Guizhou Oversea Talent Project (Qianren Xiangmu Zizhu Hetong 2004#02), and Key Project of Students in Guizhou Normal University (GNU-SP-2005-SJXY-A01)     

Evaluating landraces of bread wheat <Emphasis Type="Italic">Triticum aestivum</Emphasis> L. for tolerance to aluminium under low pH conditions

Bread wheat (Triticum aestivum L.) landraces held within ex situ collections offer a valuable and largely unexplored genetic resource for wheat improvement programs. To maximise full utilisation of such collections the evaluation of landrace accessions for traits of interest is required. In this study, 250 accessions from 21 countries were screened sequentially for tolerance to aluminium (Al) using haematoxylin staining of root tips and by root regrowth measurement. The staining test indicated tolerance in 35 accessions, with an intermediate response to Al exhibited in a further 21 accessions. Of the 35 accessions classified as tolerant, 33 also exhibited increased root length following exposure to Al. The tolerant genotypes originated from Bulgaria, Croatia, India, Italy, Nepal, Spain, Tunisia, and Turkey. AFLP analysis of the 35 tolerant accessions indicated that these represent diverse genetic backgrounds. These accessions form a valuable set of germplasm for the study of Al tolerance and may be of benefit to breeding programs for expanding the diversity of the gene pool from which tolerant cultivars are developed.     

RAPD Polymorphism of the White-Flowered Gourd (Lagenaria siceraria (Molina) Standl. Landraces and its Wild Relatives in Kenya

The present study was performed to investigate genetic diversity of Kenyan landraces of the white-flowered gourd (Lagenaria siceraria), which exhibits tremendous morphological variation. RAPD analyses were performed on 53 landraces of the cultivated species L. siceraria and 42 accessions of three wild species (40 L. sphaerica, 1 L. abyssinica, and 1 L. breviflora). A total of 432 polymorphic bands were detected using 54 primers. The four species were clearly differentiated from one another. Intra-specific variations were investigated with L. siceraria and its wild relative L. sphaerica. Landraces of the cultivated species collected from different ethnic communities or regions were differentiated. Morphological variations were not associated with RAPD variations. Bitter landraces collected in Maasai communities showed two specific RAPD bands. In the wild species, accessions collected from the eastern and western sides of the Great Rift Valley were genetically differentiated from each other. In both species, genetic and geographical distance matrices computed among all pairs of accessions were significantly correlated, implying that the observed geographical variation can be explained by the 'Isolation by distance model'. Progeny plants derived from a common mother in L. siceraria showed a low level of segregation in RAPD pattern, suggesting that collected landraces are cultivated, maintaining their inherent traits although they are monoecious and insect-pollinated, whereas the wild relative L. sphaerica showed a higher level of segregation. The morphological diversity observed among landraces of L. siceraria is the result of human selection and their genetic identities are maintained by inbreeding probably resulting from frequent self-pollination.     

Phenotypic diversity of durum wheat (<Emphasis Type="Italic">Triticum durum</Emphasis> Desf.) from Jordan

Durum wheat landrace genotypes are disappearing from the main wheat areas in Jordan, because of spreading of new uniform cultivars and the serious reduction in wheat cultivation. This study was conducted to evaluate genetic diversity in durum wheat landraces from Jordan and to identify desirable agronomic traits. Landraces were collected from two target areas: Ajloun and Karak. The collected material was grown under rainfed conditions using an augmented design with five blocks and four repeated check cultivars. Data were collected for 14 morphological and agronomic traits. Phenotypic diversity index (H′) was estimated, and the relationships among accessions were measured using cluster analysis and dendrogram similarity matrix. The results revealed the presence of a wide range of variability among landraces., which possess high levels of variability for biological yield, fertile tillers, number of seeds per spike, seed weight per spike and weight of 1000 seeds. These landraces must be considered as a reservoir of genes that plant breeders need in their wheat improvement programs and should be conserved both ex situ and in situ.     

Characterization of a flint maize (Zea mays L. convar. mays) Italian landrace: I. Morpho-phenological and agronomic traits

Maize (Zea mays L.) landraces have the highest genetic variation and adaptation to the natural and anthropological environment where they have evolved. Surveying both qualitative and quantitative morphological traits of existing landraces may be useful in maintaining their genetic diversity and preserving them from genetic erosion. Our research deals with the morpho-phenological and agronomic characterization of a flint maize landrace, named 'Nostrano di Storo', still grown in an inland hilly environment in the low valley of Chiese River in Trentino, North-Eastern Italy. The majority of plants from twenty field populations proved to belong, with few exceptions (NSt2, NSt9, NSt11), to a single population. It means that the plant material long grown in this area and maintained by local farmers through yearly selection forms a single landrace within which some populations (i.e. NSt1, NSt3, NSt4, NSt7, NSt10, NSt18, NSt19, NSt20) could be considered as most representative and taken as 'core'. This is supported by the fact that the genetic variability was much higher within than between field populations: half of the plant and ear traits investigated did not show any significant difference between populations whereas all traits but two showed highly significant differences within populations. Selection carried out over the years by each farmer according to his own criteria produced little genetic differentiation within the original population. Gene flow among farmer populations, most likely occurred through both pollen dispersion to neighboring cultivated fields and seed exchange among farmers, may help to explain the low genetic differentiation. This information is useful for both planning conservation and recognizing the landrace as a unique germplasm source of specific geographic origin.     

Genetic Diversity Among Brazilian Cultivars and Landraces of Tomato Lycopersicon Esculentum Mill. Revealed by RAPD Markers

Random amplified polymorphic DNA markers (RAPD) were used to estimate the variability of 35 tomato accessions (Lycopersicon esculentum Mill.). A total of 257 reproducibly scorable bands were obtained from 20 primers, 78.6% of which were polymorphic. The percentage distribution of RAPD markers shows a bimodal distribution, and the frequency of rare alleles is similar in commercial and landrace accessions. Genetic distances among accessions were calculated and a dendrogram showing the genetic relationships among them was constructed allowing for the separation of four groups. Twenty out of 23 Brazilian landraces fell within one group, whereas commercial cultivars were distributed in the four groups. AMOVA analysis of RAPD data showed that, despite the high within Brazilian landraces and commercial cultivars variation, these two groups are significantly different, indicating that landraces can be a source of variation for breeding programs.     

Landraces of cowpea, <Emphasis Type="Italic">Vigna</Emphasis><Emphasis Type="Italic">unguiculata</Emphasis> (L.) Walp., as potential sources of genes for unique characters in breeding

Cowpea landraces belonging to Vigna unguiculata (L.) Walp. subsp. unguiculata cv.-gr. unguiculata and cv.-gr. sesquipedalis collected from part of the Deccan Plateau and West Coast of India were evaluated to (i) identify the diverse source(s) of variation for improved characters like pods/peduncle and seed index (ii) study the response of landraces for adaptation to drought and heat stress and (iii) understand the breeding value of a landrace in the genetic improvement of a popular cowpea cultivar. Landraces were evaluated for various morphological characters, pods/peduncle, seed index and other economically important agronomic traits, rust resistance and drought and heat tolerance in different years and environments. Landraces were found as an important source of genetic variability for pods/plant, pods/peduncle, better pod filling ability (seed index), grain yield/plant as well as drought and heat tolerance and rust resistance. Hybridization between C 152 (cv.) and DWDCC 016 (landrace) resulted in release of new variation not present in the two parents. Thus the landrace, DWDCC 016, can be utilized to improve cultivated varieties by transferring to them the economically valuable traits like pods/peduncle and seed index thereby enhancing realisation of sink potential and ultimately grain yield in a sustainable way.     

Phenotypic Variation and Relationships in Landraces and Improved Varieties of Rye (Secale cereale L.) from Northern Europe

Phenotypic diversity for agronomic characteristics was determined in an experiment with 29 landraces and 14 improved varieties of rye from the Nordic area, Germany and Poland. The accessions were scored for 12 characters. Effects of the location, year, type, and country of origin for landraces were investigated with analysis of variance. Phenotypic variations for the traits were estimated using the Shannon–Weaver diversity index. The genetic variation was high, with an average of H ₀ = 0.566. The landraces from Norway, Sweden and Finland showed the highest variation, whereas the improved varieties had the lowest. The German material also had low variation but the status of this material is uncertain. The genetic diversity showed that 70% of the variation was found within the accessions. A cluster analysis was carried out to identify the relationship between the accessions. The material grouped into eight clusters, where clusters I to V included landraces from Sweden, Finland and Norway, except for cluster III, which included one improved variety from Denmark. Cluster VI comprised a single Swedish landrace from Gotland. Most of the improved varieties were in cluster VII and the last cluster contained accessions from Germany and Sweden.     

Diversity of Nordic Landrace Potatoes (<Emphasis Type="Italic">Solanum tuberosum</Emphasis> L.) Revealed by AFLPs and Morphological Characters

The management of genebank collections of cultivated potato is costly due to the need for in vitro maintenance and virus eradication. Therefore, it is important to set up conservation strategies, which prevent duplicates entering the collections. In this study, 32 Nordic potato landraces were studied for 57 morphological traits and analysed for amplified fragment length polymorphism (AFLP). Most landraces could be distinguished based on the morphological characters, except five accessions. Using five primer combinations to generate 114 reproducible AFLPs, of which 63 (55%) were polymorphic, the five morphologically indistinguishable accessions were placed into two groups with identical AFLP patterns, suggesting that some of the accessions were redundant for long-term preservation. The AFLP data showed that the Nordic collection of potato landraces is composed of genetically different clones, and morphological analysis revealed a wide range of variability. This variability seems to be distributed randomly over the Nordic region since the cluster analysis based on AFLPs and morphological traits revealed no grouping based on the country of origin. Principal component analysis suggests that fewer morphological traits than used in this study will be sufficient to discriminate between different genotypes of cultivated potato (Solanum tuberosum L.). Future possibilities for rationalising potato collections are discussed.     

Molecular diversity of Omani wheat revealed by microsatellites: I. Tetraploid landraces

Results of archaeological studies indicate a millennia-old cultivation history for wheat (Triticum spp.) in Oman. However, in spite of numerous collection surveys and efforts for phenotypic characterization of Omani wheat landraces, no attempts have been made using molecular tools to characterize this germplasm. To fill this gap, 29 microsatellite markers revealing 30 loci were used to study the genetic diversity of 38 tetraploid wheat landrace accessions comprising the species T. dicoccon, T. durum and T. aethiopicum. A total of 219 alleles were detected whereby the number of alleles per locus ranged from 2 to 16 with an average number of 7.1 alleles per locus. The highest number of alleles occurred in the B genome with on average 7.9 alleles per locus as compared to the A genome with 6.5 alleles per locus. Heterogeneity was detected for all microsatellites except for GWM 312, GWM 601 and GWM 192B with an average heterogeneity over all primers and lines of 14.4%. Approximately 10% of the accessions contained rare alleles with an average allele frequency <4%. Gene diversity across microsatellite loci ranged from 0.26 to 0.85. The pairwise comparison of genetic similarity ranged from 0.03 to 0.91 with an average of 0.2. Cluster analysis revealed a clear separation of the two species groups T. dicoccon versus T. durum and T. aethiopicum. Within the species clusters regional patterns of subclustering were observed. Overall, this study confirmed the existence of a surprisingly high amount of genetic diversity in Omani wheat landraces as already concluded from previous morphological analyses and showed that SSR markers can be used for landraces’ analysis and a more detailed diversity evaluation.     

Genetic Diversity and Relationships of Wheat Landraces from Oman Investigated with SSR Markers

Little is known about genetic diversity and geographic origin of wheat landraces from Oman, an ancient area of wheat cultivation. The objectives of this study were to investigate the genetic relationships and levels of diversity of six wheat landraces collected in Oman with a set of 30 evenly distributed SSR markers. The total gene diversity, (H_T), conserved in the three durum wheat (Triticum durum desf.) landraces (H_T = 0.46) was higher than in the three bread wheat (Triticum aestivum L.) landraces (H_T = 0.37), which were similar to Turkish and Mexican bread wheat landraces calculated in previous studies. Genetic variation partitioning (G_ST) showed that variation was mainly distributed within rather than among the durum (G_ST = 0.30) and bread wheat (G_ST = 0.19) landraces. Based on modified Rogers’ distance (MRD), the durum and bread wheat landraces were distinct from each other except for a few individuals according to principal coordinate analysis (PCoA). One bread wheat landrace (Greda) was separated into two distinct sub-populations. A joint cluster analysis with other landraces of worldwide origin revealed that Omani bread wheat landraces were different from other landraces. However, two landraces from Pakistan were grouped somewhat closer to Omani landraces indicating a possible, previously unknown relationship. Implications of these results for future wheat landrace collection, evaluation and conservation are discussed.     

Genetic diversity of landraces of wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) from hilly areas of Uttaranchal,India

Seed samples of 27 landraces of wheat were collected from farmers’ fields of hilly areas of Himalaya in Uttaranchal state of India during April 2004. Genetic diversity among 41 genotypes (cultivars and landraces of wheat) was studied using morphological traits, microsatellite markers and SDS-PAGE of HMW-GS. The dendrogram and PCA (Principal Component Analysis) based on morphological data clearly separated landraces of wheat from cultivars. In the dendrogram based on microsatellite markers data all the wheat cultivars released after the introduction of high yielding dwarf wheat varieties from CIMMYT, used in this study, were grouped separately with the exception of NP4. The pre-green revolution indigenous varieties grouped with landraces suggesting that the same had been probably developed through selection among landraces in India. The landraces had higher diversity for HMW-glutenin subunits coded by Glu-B1, with distinct subunit combinations 6 + 8, 7 + 9, 13 + 16, than within the wheat cultivars analyzed. Most of the landraces except IITR10 and IITR14 are clearly distinct from the indigenous and modern wheat cultivars released in India in the 20th century. More than half of the landraces were heterogeneous mixture of plants with different glume color, awnness, grain color and HMW-GS profile and hence need purification through single plant selection. Some of the landraces with resistance to yellow rust and powdery mildew and distinct HMW-GS subunits can be used in appropriate breeding programs. It will be desirable to conserve and protect the landraces as geographical indications of Uttaranchal.     

Evaluation of genetic diversity and relationships within an on-farm collection of Perilla frutescens (L.) Britt. using microsatellite markers

The present study demonstrates utilization of 11 microsatellite markers to explore genetic diversity held in Perilla frutescens (L.) Britt. landrace accessions growing on farms in different parts of Korea and Japan and to assess their genetic relationships. All microsatellite loci were polymorphic and produced a total of 96 alleles ranging from 4 to 20, with an average of 8.7 alleles per locus. Of the 96 alleles found, a total of 15 unique landrace-specific alleles were observed at 9 different loci. The locus GBPFM203 provided the highest number of alleles (20), of which five were unique and each specific to a particular landrace accession. The occurrence of unique, accession-specific alleles presented molecular evidence for the generation of new alleles within on-farm collection of Perilla. The mean values of observed (H _O) and expected heterozygosity (H _E) were 0.39 and 0.68, respectively, indicating a considerable amount of polymorphism within this collection. A genetic distance-based phylogeny grouped the two Perilla varieties, var. frutescens and var. crispa (Thunb.) Decne into two distinct groups. Accessions belonging to var. frutescens could also be divided into two subgroups at a close genetic distance (GD = 0.432). The overall clustering pattern did not strictly follow the grouping of accessions according to their geographic origins. These observations are indicative of extensive germplasm exchange among farms from different geographical regions. The genetic similarity observed among the Perilla landraces may be useful for future Perilla crop variety identification, conservation, and improvement programs.     

Different cucumber melon (<Emphasis Type="Italic">Cucumis melo</Emphasis> L.) races cultivated in Salento (Italy)

South Italy is still a centre of diversity for melons but new cultivars progressively replace its traditional landraces. The area of Salento is important for several typical old crops as the cucumber melon ‘Meloncella’ (Cucumis melo L. var. chate (Hasselq.) Filov) traditionally cultivated for its unripe fruits. Information on cultivation, quality and variation of this Apulian landrace (probably the last relic of a wider cultivation in Europe) is reported. A strategy for its characterization and safeguarding is in progress at the genebank of IGV of Bari (Italy).     

Can Phaseolin Patterns Help Resolve the Phaseolus–Vigna Complex?

Although common bean (Phaseolus vulgaris L.) is cultivated throughout India, the Himalayas hold largest diversity of bean germplasm. No studies on characterization of phaseolin types on this germplasm have been conducted earlier. In order to determine whether the common bean cultivars collected from various areas in the Northern Himalayas represent introductions from the Central American and Andean domestication centers or are local domesticates, we have analyzed the electrophoretic variation (SDS-PAGE) of phaseolin types in several bean accessions. A few species of Vigna were also included in this study to determine whether phaseolin (vignin in Vigna) patterns can be used to resolve the Phaseolus–Vigna complex. The present investigation on phaseolin (globulin) patterns of Phaseolus vulgaris and Vigna spp. clearly shows much variability in globulin patterns. Three new types of phaseolin patterns were recorded. An attempt to resolve phylogenetic problems in this complex was made using the phaseolin data.     

Determining Sources of Diversity in Cultivated and Wild <Emphasis Type="Italic">Lablab purpureus</Emphasis> Related to Provenance of Germplasm by using Amplified Fragment Length Polymorphism

To improve understanding of diversity of Lablab purpureus and establish relationships among 103 germplasm accessions collected from diverse geographic origins, amplified fragment length polymorphism markers were used. Four primer sets selected out of 16 produced 289 clear, repeatable polymorphisms. UPGMA analysis of similarity data clustered the accessions according to their subspecific taxonomic organization, i.e., subsp. purpureus and subsp. uncinatus, as well as to cultivated and wild forms. The well-represented landraces from Africa and Asia, belonging predominantly to subsp. purpureus, displayed moderate genetic diversity. Wild forms from Africa showed far greater levels of diversity that would justify taxonomic re-assessment of the wild subsp. uncinatus. The molecular analysis identified forms that were collected in the wild in India but were genetically placed intermediate between wild and cultivated forms. As these plant types did not exist among the African accessions, it is suggested that they might represent escapes from early attempts of domestication. These results support the suggested pathway of domestication and distribution of L. purpureus from Africa to Asia. Additional members to a previously published core collection of the species are proposed.