Analysis of Genetic Diversity Among Saccharum spontaneum L. From Four Geographical Regions of India, Using Molecular Markers

Saccharum spontaneum L. a wild relative of sugarcane is the most variable and diverse among the Saccharum species. This species had been successfully exploited in sugarcane improvement programmes since 1915 and most of the present day commercial varieties are derivatives of interspecific hybrids involving S. spontaneum. The S. spontaneum germplasm available today in the World collections is diverse and represent different geographical groups. In the present investigation, an attempt was made to characterize 40 S. spontaneum clones collected from 4 different geographical areas in India using 20 random, 2 ISSR and 2 telomere primers. Of the 491 bands generated by these primers 83.9% were polymorphic. The genetic diversity estimated based on these markers was found to be moderate (48.9%). The pair-wise genetic distance between the accessions ranged from 29.8 to 60.0. The accessions from Arunachal Pradesh were found to be the most diverse, while Tamil Nadu accessions showed relatively less diversity. Diversity between Tamil Nadu and Kerala collections was found to be low, while the diversity between the Orissa group and the rest was found to be high. The collections from Mayurbanj and Cuttack regions of Orissa were found to be distinct. Arunachal Pradesh accessions, being more diverse, are a potential source for exploitation in sugarcane breeding programmes.     

Marker-Assisted Acquisition and Core Collection Formation: A Case Study in Barley using AFLPs and Pedigree Data

A problem that often occurs in deciding which germplasm should be acquired to expand the diversity of a plant genetic resources collection, and which accessions should be included in a core collection, is the lack of proper data. The usefulness of an AFLP-based protocol to assist in acquisition decisions and in core collection formation was examined by using 52 barley cultivars. For validation purposes, pedigree data of the cultivars were used to calculate the ‘effective number of origin lines’ (n_o1), a parameter introduced in earlier research that was defined as the number of alleles per locus, not identical by descent, in a set of lines. Two AFLP primer combinations were able to distinguish all 52 cultivars from each other, and to discriminate between spring and winter crop types. Using the year of origin of the cultivars, the historical development of n_o1 showed a stepwise pattern, indicating the periodical release of genetically similar cultivars, alternated by the incorporation of new material. Comparison of AFLP data between cultivars and both their parents was possible in five cases. These comparisons revealed a high likelihood that the correct parents were involved but a rather skewed contribution of parents to offspring, suggesting that backcrossing had been applied. Treating the 25 cultivars that were released before 1980 and played an important role in barley cultivation as a basic collection, and the 27 more recent cultivars as potential candidates for acquisition, n_o1 values generated by a marker-based approach largely followed those using a random approach. Given this poor performance, a marker-based protocol to assist in acquisition decisions was not considered useful for the analysed material. If the 52 cultivars were considered to be the collection from which a core collection had to be selected, the marker-based selection showed much better results compared to a random selection. About half of the total number of origin lines could be captured with a quarter of the collection, indicating the potential utility of AFLPs in core collection formation.     

Development of Core Collection in Pigeonpea [<Emphasis Type="Italic">Cajanus cajan</Emphasis> (L.) Millspaugh] using Geographic and Qualitative Morphological Descriptors

Pigeonpea is an important pulse crop grown by smallholder farmers in the semi-arid tropics. Most of the pigeonpea cultivars grown to date are selections from the landraces, with a narrow genetic base. With the expansion of the crop to newer areas, problems of local importance are to be addressed. Hence, an economically feasible and faster germplasm evaluation mechanism, such as a core collection, is required. This article describes the development of core collection from 12,153 pigeonpea accessions collected from 56 countries and maintained at ICRISAT, Patancheru, India. The germplasm accessions from 56 countries were placed under 14 clusters based primarily on geographic origin. Data on 14 qualitative morphological traits were used for cluster formation by Ward’s method. From each cluster ≈10% accessions were randomly selected to constitute a core collection comprising 1290 accessions. Mean comparisons using Newman–Keuls test, variances’ comparisons by Levene’s test, and comparison of frequency distribution by χ²-test indicated that the core collection was similar to that of the entire collection for various traits and the genetic variability available in the entire collection is preserved in the core collection. The Shannon–Weaver diversity index for different traits was also similar for both entire and core collection. All the important phenotypic associations between different traits available in the entire collection were preserved in the core collection. The core collection constituted in the present study facilitates identification of useful traits economically and expeditiously for use in pigeonpea improvement.     

Sampling a Core Collection of Island Cotton (Gossypium barbadense L.) Based on the Genotypic Values of Fiber Traits

A genetic model, including effects of environments, genotypes, and genotype by environment interaction, was employed to analyze five fiber traits of Island cotton (Gossypium barbadense L.). Genotypic values of 304 accessions were predicted by the adjusted unbiased prediction (AUP). Genetic similarities between different accessions were measured by Mahalanobis distances based on genotypic values. Appropriate sampling strategies, linkage rules in stepwise clustering, and sampling proportion were evaluated. To form a core collection of Island cotton, 60 accessions were sampled by the deviation sampling strategy combined with single linkage rule of hierarchical clustering. The genetic variation and structure captured by the core collection were examined in means, variances, ranges and coefficients of variation, correlation coefficients of quantitative traits, and the accessions distribution plotted by first two principal components between two collections. It was showed that the initial collection was well represented by the core collection for exploiting the Island cotton germplasm. These authors contributed equally to this work.     

An assessment of the genetic diversity within a collection of<Emphasis Type="Italic">Saccharum spontaneum</Emphasis> L. with RAPD-PCR

A local collection of 33Saccharum spontaneum L. clones and two sugarcane cultivars (LCP 82-89 and LCP 85-384) were assessed for genetic variability using random amplified polymorphic DNA (RAPD)-PCR. A total of 157 polymorphic RAPD-PCR bands were scored with 17 primers. The number of RAPD-PCR products per primer ranged from four to 16. The data were analyzed with two multivariate analysis software programs, NTSYSpc and DNAMAN®. Although these two programs yielded similar results, a bootstrapped phylogenetic tree could only be generated with the DNAMAN® software. A substantial degree of genetic diversity was found within the localS. spontaneum collection. Pairwise genetic homology coefficients ranged from 65% (SES, 196/Tainan 2n = 96) to 88.5% (IND 81-80/IND 81-144). LCP 82-89 and LCP 85-384 shared a greater similarity (82%) than either was to any clone ofS. spontaneum (ranging from 60.5 to 75.2%). The 33S. spontaneum clones were assigned to eight groups independent of their geographic origin or morphology, while the two sugarcane cultivars were assigned to the ninth group. All but two pairs ofS. spontaneum clones could be distinguished by a single RAPD primer OPBB-02. The use of a second primer, either OPBE-04 or Primer 262, separated allS. spontaneum clones. One amplification product from the RAPD primer OPA-11, OPA-11-336, proved to be cultivar-specific and has been adopted for use in our breeding program. Information from this study would help conserve the genetic diversity ofS. spontaneum. Disclaimer: Product names and trademarks are mentioned to report on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by USDA does not imply the approval of the product to the exclusion of others that may also be suitable. The experiments reported comply with the current laws of the USA.     

Collection of Lupinus angustifolius L. Germplasm and Characterisation of Morphological and Molecular Diversity

Lupinus angustifolius L. is a Mediterranean species, domesticated in the 20th century, representing an important grain legume crop in Australia and other countries. This work is focused on the collection of wild germplasm and on the characterisation of morphological and molecular diversity of germplasm accessions. It reports the collection of 81 wild L. angustifolius accessions from the South and Centre of Portugal, available at the ‘Instituto Superior de Agronomia Gene Bank’, with subsequent morphological and molecular characterisation of a selection of these and other accessions. A multivariate analysis of morphological traits on 88 L. angustifolius accessions (including 59 wild Portuguese accessions, 15 cultivars and 14 breeding lines) showed a cline of variation on wild germplasm, with plants from Southern Portugal characterised by earlier flowering, higher vegetative development and larger seeds. AFLP and ISSR molecular markers grouped modern cultivars as sub-clusters within the wider diversity of wild germplasm, revealing the narrow pool of genetic diversity on which domesticated accessions are based. The importance of preserving, characterising and using wild genetic resources for L. angustifolius crop improvement is outlined by the results obtained.     

Characterization of a USDA Kentucky Bluegrass (Poa pratensis L.) Core Collection for Reproductive Mode and DNA Content by Flow Cytometry

Kentucky bluegrass (Poa pratensis L.) is an important turf and forage grass species with a facultative apomictic breeding behavior. In this study, mature seed and leaf tissue from 38 accessions of a USDA core collection of Kentucky bluegrass were analyzed with flow cytometry to characterize the reproductive mode and DNA content for each accession. Major reproductive pathways for each accession were determined based upon the presence and the position of the peaks observed and the known methods of reproduction for Kentucky bluegrass. While the majority of the accessions exhibited facultative apomictic reproductive behavior with a combination of reduced, zygotic and unreduced, parthenogenic embryo production, obligate sexual or obligate apomictic accessions were also found to be present in this core collection. In addition, reduced, parthenogenic and unreduced zygotic embryos were also detected in several accessions. Flow cytometric analysis of somatic tissue revealed a large range of DNA variation within this core collection. We also examined the sensitivity of flow cytometry in analyzing bulked samples containing a large number of plants with varied DNA content and determined that flow cytometry can effectively detect a plant having a different DNA content within a 15-plant bulk sample. Overall the combination of mature seed and somatic tissue analysis generated important information for the Kentucky bluegrass core collection and can be an effective and affordable tool to characterize even greater numbers of Kentucky bluegrass accessions.     

Analysis of the contribution of Mesoamerican and Andean gene pools to European common bean (<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis> L.) germplasm and strategies to establish a core collection

Common bean (Phaseolus vulgaris L.) was introduced in Europe from both Mesoamerican and Andean centres of origin. In this study, a collection including 544 accessions from all European regions showed that the Andean phaseolin types ‘T’ (45.6%) and ‘C’ (30.7%) prevailed over the Mesoamerican ones ‘S’ (23.7%), and accessions with cuboid seed shape (34.9%), maroon coat darker colour seed (44.3%), uniform seed colour (69.6%) were the most frequent. European accessions with phaseolin ‘S’ showed a significantly larger average seed size compared to those from America in the same phaseolin class while those presenting ‘T’ and ‘C’ phaseolin did not. This suggests that, during crop expansion in Europe, sampling or selection favoured the large-seeded races within the Mesoamerican ‘S’ gene pool or, possibly, introgression from Andean germplasm did occur. A core collection was developed using sampling approaches based on the information available in the genebank databases and on phaseolin patterns. Four sampling strategies were used: simple random sampling, and three random-stratified samplings, by logarithm of frequency of accessions by country, by European region, and by phaseolin pattern, respectively. Two sampling strategies resulted in core collections significantly different for phaseolin electrophoretic patterns from the whole collection. Stratification by phaseolin patterns increased the frequency of ‘S’ types (‘C’ type = 33%, ‘T’ type = 5.7% and ‘S’ type = 31.3%). The core collections were validated using seven seed characters, and no significant difference was observed in all strategies. This first developed European bean core collection will help to assess the contribution of the two American gene pools to the European germplasm and their relative importance for breeding purposes.     

Phenotypic diversity and morphological characterization of Amygdalus L. species in Lebanon

In this paper, we report on morphological diversity of Amygdalus L. species native to Lebanon, following a countrywide survey of almond germplasm whereby a total of 149 accessions were collected throughout the country and characterized by thirteen quantitative and four qualitative traits. The results indicated that the genetic diversity of Amygdalus communis L., Amygdalus korshinskyi Hand.-Mazz., and Amygdalus orientalis Duh. in Lebanon is high. Principal component analysis revealed that nut weight, nut volume, nut width, kernel volume and shell strength had highest loading in the first component that accounted for 38.7% and 46.7% of total variation in A. communis and A. orientalis, respectively. In contrast, leaf traits were present in the second component which accounted for 18% and 23.2% of total variation in each species, respectively. No significant correlations were detected between leaf parameters and fruit traits in both species. The results indicated that quantitative leaf characters for all three species were determined by rainfall and not altitude whereby adjacent accessions located in drier areas had smaller leaf sizes than those located in more humid regions. Quantitative fruit characters did not seem to vary accordingly. Qualitative leaf traits in all three species reflected a variability which was independent of rainfall. A. communis populations showed high variability, suggesting that they could be a valuable source in almond improvement programs.     

A method for establishing a core collection of Saccharum officinarum L. germplasm based on quantitative-morphological data

A technique is proposed for establishing a representative core collection of S. officinarum accessions from the world collection of sugarcane germplasm maintained at the Sugarcane Breeding Institute-Research Centre, Cannanore, India. In the proposed method, the accessions were first sorted based on their relative contributions to the total variability by means of principal component scores based on a set of quantitative characters. Then, the cumulative proportion of their contributions to the total variance was computed. A logistic regression model was fitted to evaluate the functional relationship between the cumulative proportion of variance and the number of accessions. The size of the core set was decided as the inflection point on that fitted curve, i.e., the point beyond which the rate of increase in cumulative proportion of variability contributed by an accession began to decline. A method for eliminating entries with a high degree of similarity from the selected core set is also proposed.     

Patterns of Phenotypic Variation in a Germplasm Collection of Carthamus tinctorius L. from the Middle East

Phenotypic diversity was assessed for quantitative and qualitative traits in a salt-tolerant subset of the international safflower (Carthamus tinctorius L.) germplasm collection originating from 11 countries in three regions (Central Asia, Southwest Asia and Africa) of the Middle East. Phenotypically, the germplasm, among and within regions, was highly variable, especially for rosette- and yield-related traits. Frequency of desirable variants of seven agronomically important traits ranged from 14% for long rosette period to 50% for no or few spines. Level of population differentiation was high for number of capitula per plant (30%), whereas most traits partitioned their diversity (82–87%) within populations. Region-specific nonrandom associations among sets of qualitative traits and the existence of broad morphological and phenotypic diversity in this germplasm were supported by the large number of log-linear models needed to describe qualitative trait associations, the high number of principal components needed to account for total variability, and the low discriminatory power of phenotypic traits among germplasm from regions and countries in the Middle East. These results suggest that adaptation of the species to the wide spatial and temporal variation in the Middle East resulted in a multitude of ecotypes and in enormous amount of local variation. A multivariate selection criterion for high biological and seed yield, long rosette period and no or few spines identified five accessions from Southwest Asia that can be introduced into subsistence farming systems as a multipurpose crop under saline agriculture.     

Development of Core Subset of Finger Millet Germplasm Using Geographical Origin and Data on 14 Quantitative Traits

Finger millet [Eleusine coracana (L.) Gaertn.] is an important cereal food crop in Africa and South Asia. It is a hardy crop that can be grown in very diverse environments from almost at sea level to about 2400 m.a.s.l. Finger millet has an excellent food value as its seeds contain protein ranging from 7 to 14% and are particularly rich in methionine amino acid, iron, and calcium. Despite all these merits, this crop has been neglected from the main stream of crop improvement research. One of the means to boost its production and productivity is to enhance utilization of finger millet germplasm to breed superior varieties. Keeping this objective in view, a core subset of finger millet germplasm (622 accessions) based on origin and data on 14 quantitative traits was developed from the entire global collection of 5940 accessions held in the genebank at ICRISAT, Patancheru, India. The comparison of means, variances, frequency distribution, Shannon-Weaver diversity index (H′) and phenotypic correlations indicated that the core subset represents the entire collection. These tests indicated that sampling was optimal and the diversity has been captured very well in the core subset. The correlation analysis indicated that panicle exsertion and longest finger length could be given lower priority in the future germplasm evaluation work of finger millet.     

Collection and Characterization of Citrus indica Tanaka and C. macroptera Montr.: Wild Endangered Species of Northeastern India

Citrus indica and C. macroptera are the wild endangered species of Citrus occurring in northeastern India. Surveys were undertaken in this region for ascertaining distribution, studying variability and for collection of germplasm of these two species. C. indica, an endemic species of this region, was collected from the Citrus Gene Sanctuary located in buffer zone of Nokrek Biosphere Reserve in the Garo hills of Meghalaya. In addition, a putative natural hybrid of C. indica and C. limon was collected for the first time from the south Garo hills. C. macroptera had much wider distribution and was collected from Mizoram and Meghalaya states. In Jantia hills of Meghalaya, natural populations of this species are in a highly threatened state. The two species were unevenly distributed all over the explored territory. Morphological characterization of leaves, fruits and seeds indicated the presence of sizable variability within collected accessions of these two Citrus species. Indigenous technical knowledge gathered on the use and socio-economic importance indicated commercial potential for these two species in northeastern India. However, lack of cultivation of these species and clearing of forest cover at an alarming rate has led to an urgent need to adopt complementary conservation strategies to safeguard these species and to ensure their availability for future utilization. A major emphasis on developing methods for their propagation, multiplication and regeneration in in situ and ex situ conditions is required.     

Sampling effects on the assessment of genetic diversity of rhizobia associated with soybean and common bean

Biological nitrogen fixation plays a key role in agriculture sustainability, and assessment of rhizobial diversity contributes to worldwide knowledge of biodiversity of soil microorganisms, to the usefulness of rhizobial collections and to the establishment of long-term strategies aimed at increasing contributions of legume-fixed N to agriculture. Although in recent decades the use of molecular techniques has contributed greatly to enhancing knowledge of rhizobial diversity, concerns remain over simple issues such as the effects of sampling on estimates of diversity. In this study, rhizobia were isolated from nodules of plants grown under field conditions, in pots containing soil, or in Leonard jars receiving a 10⁻² or a 10⁻⁴ serially-diluted soil inoculum, using one exotic (soybean, Glycine max) and one indigenous (common bean, Phaseolus vulgaris) legume species. The experiments were performed using an oxisol with a high population (10⁵ cells g⁻¹ soil) of both soybean rhizobia, composed of naturalized strains introduced in inoculants and of indigenous common-bean rhizobia. BOX-PCR was used to evaluate strain diversity, while RFLP-PCR of the ITS (internally transcribed spacer) region with five restriction enzymes aimed at discriminating rhizobial species. In both analyses the genetic diversity of common-bean rhizobia was greater than that of soybean. For the common bean, diversity was greatly enhanced at the 10⁻⁴ dilution, while for the soybean dilution decreased diversity. Qualitative differences were also observed, as the DNA profiles differed for each treatment in both host plants. Differences obtained can be attributed to dissimilarity in the history of the introduction of both the host plant and the rhizobia (exotic vs. indigenous), to host-plant specificity, rhizobial competitiveness, and population structure, including ease with which some types are released from microcolonies in soil. Therefore, sampling method should be considered both in the interpretation and comparison of the results obtained in different studies, and in the setting of the goals of any study, e.g. selection of competitive strains, or collection of a larger spectrum of rhizobia. Furthermore, effects of sampling should be investigated for each symbiosis.     

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.     

Yield Potential and Ecophysiological Traits of the Altamurano Linseed (Linum usitatissimum L.), a Landrace of Southern Italy

Altamurano is a linseed landrace of southern Italy. Its area of origin is characterised by typical Mediterranean climate. Traditionally, Altamurano is spring sown, and mainly used as a feed integrator for animals. This landrace was evaluated during 2 years within varietal (16 modern varieties) and sowing date (15 sowings, from October to April, with ‘Antares’ as a control) trials. Growth analysis was carried out on two sowings. Seed yield was slightly lower than modern varieties, with advantages in late sowings. Reproductive efficiency, mean seed weight, harvest index and oil percentage were often higher than in modern cultivars. Altamurano was also characterised by early flowering, low vegetative growth, long leaf area duration, high light use efficiency during early grain filling phase, and relatively long grain filling period. All these features are connected with its adaptation to spring sowings in Mediterranean areas, characterised by early spring drought. Despite the decrease of its cultivation area from about 400 to less than 100 ha in the last 10 years, Altamurano is still present in the more traditional farms of the area as a heritage of past growing systems and historical adaptation to the environment.     

Genetic diversity of the African hexaploid species Solanum scabrum Mill. and Solanum nigrum L. (Solanaceae)

Two hexaploid species of Solanum sect. Solanum are present in Africa: Solanum scabrum and S. nigrum. Solanum scabrum is a widely cultivated species and is used as a leafy vegetable, as a source of medicine and as a source of ink dye. In previous studies a wide range of morphological diversity has been reported in this species and in some studies subspecies have been proposed. Subspecies are also recognized in S. nigrum. However, it has not been established whether or not the morphological differences are reflected at the genomic level. The present study applies AFLPs to study the genetic diversity in S. scabrum and its relationship to geographical provenance, morphological differences and the possible existence of subspecies within S. scabrum and S. nigrum. The data obtained were analyzed with cluster analysis (using UPGMA and NJ). The results indicate that the genetic variation within S. scabrum was higher within accessions than between accessions. Accessions did not cluster according to their geographical provenance, indicating that accessions from different geographical areas were not significantly different genetically. The clustering reflected neither morphological differences nor domestication status (cultivated or wild). The morphological differences exhibited by S. scabrum could be due to selection by farmers for different plant types. The AFLP derived clustering pattern did not segregate the subspecies recognized in S. scabrum and S. nigrum into separate subclusters.     

Analysis of Genetic Variation Among Accessions of <Emphasis Type="Italic">Pueraria montana</Emphasis> (Lour.) Merr. var. <Emphasis Type="Italic">lobata</Emphasis> and <Emphasis Type="Italic">Pueraria phaseoloides</Emphasis> (Roxb.) Benth. based on RAPD Markers

Pueraria montana var. lobata and P. phaseoloides originating from tropical Asia and parts of Oceania are ecologically and economically important legumes that are used as green manure, cover crop or forage plants. Conservation and use of plant genetic resources require an understanding of the extent and distribution of genetic diversity in any given region. In this study, genetic variation of five P. montana var. lobata and 16 P. phaseoloides accessions was analysed developing a random amplified polymorphic DNA (RAPD) marker methodology for Pueraria species and thereby creating basic data for follow-up research and the development of conservation strategies. Seeds were collected from native populations in Bac Kan Province, a mountainous region in Northeast Vietnam. P. montana var. lobata presented a high level of variation with 54.3% of the detected markers being polymorphic, whereas P. phaseoloides exhibited an intermediate to high level of variation (45.5%). The P. montana var. lobata accessions clustered in congruence with their eco-geographical origin. For P. phaseoloides no correspondence between sampling sites and genetic differentiation was found. Inter-population differentiation was measured as Jaccard's similarity coefficient (JSC). Mean JSC amounted to 0.35 in P. montana var. lobata and 0.52 in P. phaseoloides. Results are compared to other genetic studies of herbaceous legumes and conservation strategies are suggested.