Assessment of Genetic Diversity in Three Subsets Constituted from the ICRISAT Sorghum Collection Using Random vs Non-Random Sampling Procedures B. Using molecular markers

The large size of the sorghum [Sorghum bi-color (L.) Moench] landrace collection maintained by ICRISAT lead to the establishment of a core collection. Thus, three subsets of around 200 accessions were established from: (1) a random sampling after stratification of the entire landrace collection (L), (2) a selective sampling based on quantitative characters (PCS), and (3) a selection based on the geographical origin of landraces and the traits under farmers’ selection (T). An assessment was done of the genetic diversity retained by each sampling strategy using the polymorphisms at 15 microsatellite loci. The landraces of each subset were genotyped with three multiplex polymerase chain reactions (PCRs) of five fluorescent primer-pairs each with semi-automated allele sizing. The average allelic richness for each subset was equivalent (16.1, 16.3 and 15.4 alleles per locus for the subsets PCS, L, and T, respectively). The average genetic diversity was also comparable for the three subsets (0.81, 0.77 and 0.80 for the subsets PCS, L, and T, respectively). Allelic frequency distribution for each subset was compared with a chi-square test but few significant differences were observed. A high percentage of rare alleles (71 to 76% of 206 total rare alleles) was maintained in the three subsets. The global molecular diversity retained in each subset was not affected by a sampling procedure based upon phenotypic characters.     

Assessment of genetic diversity within and between pearl millet landraces

A minimum core subset of pearl millet [Pennisetum glaucum (L.) R. Br.], which comprised 504 landrace accessions, was recently established from the global pearl millet germplasm collection of ICRISAT. The accessions for this core were selected by a random proportional sampling strategy following stratification of the entire landrace collection (about 16,000 accessions) according to their geographic origin and morpho-agronomic traits. In this study RFLP probes were used to quantify the genetic diversity within and between landrace accessions of this minimum core using a subset comprising ten accessions of Indian origin. Twenty five plants per accession were assayed with EcoRI, EcoRV, HindIII and DraI restriction enzymes, and 16 highly polymorphic RFLP probes, nine associated with a quantitative trait loci (QTLs) for downy mildew resistance, and five associated with a QTL for drought tolerance. A total of 51 alleles were detected using 16 different probe-enzyme combinations. The partitioning of variance components based on the analysis of molecular variance (AMOVA) for diversity analysis revealed high within-accession variability (30.9%), but the variability between accessions was significantly higher (69.1%) than that within the accessions. A dendrogram based on the dissimilarity matrix obtained using Ward's algorithm further delineated the 250 plants into ten major clusters, each comprised of plants from a single accession (with the exception of two single plants). A similar result was found in an earlier study using morpho-agronomic traits and geographic origin. This study demonstrated the utility of RFLP markers in detecting polymorphism and estimating genetic diversity in a highly cross-pollinated species such as pearl millet. When less-tedious marker systems are available, this method could be further extended to assess the genetic diversity between and within the remaining accessions in the pearl millet core subset.     

Selective microenvironmental effects play a role in shaping genetic diversity and structure in a Phaseolus vulgaris L. landrace: implications for on-farm conservation

Little is known about the organization of landrace diversity and about the forces that shape and maintain within- and among-landrace population diversity. However, this knowledge is essential for conservation and breeding activities. The first aim of this study was to obtain some insight into how variation has been sculptured within a cultivated environment and to identify the loci that potentially underlie selective effects by using a Phaseolus vulgaris L. landrace case study whose natural and human environment and morpho-physiological traits are known in detail. The second aim of this study was to define an appropriate on-farm conservation strategy which can serve as a model for other populations. The farmers' populations of this threatened landrace were examined with 28 single sequence repeat molecular markers. The landrace appears to be a genetically structured population in which substantial diversity is maintained at the subpopulation level (62% of the total variance). Evidence of locus-specific selective effects was obtained for five of the 13 loci-differentiating subpopulations. Their role is discussed. Our data suggest that a complex interaction of factors (differential microenvironmental selection pressures by farmers and by biotic and abiotic conditions, migration rate and drift) explains the observed pattern of diversity. Appropriate on-farm conservation of a structured landrace requires the maintenance of the entire population.     

Effects of ascertainment bias and marker number on estimations of barley diversity from high-throughput SNP genotype data

The capability of molecular markers to provide information of genetic structure is influenced by their number and the way they are chosen. This study evaluates the effects of single nucleotide polymorphism (SNP) number and selection strategy on estimates of germplasm diversity and population structure for different types of barley germplasm, namely cultivar and landrace. One hundred and sixty-nine barley landraces from Syria and Jordan and 171 European barley cultivars were genotyped with 1536 SNPs. Different subsets of 384 and 96 SNPs were selected from the 1536 set, based on their ability to detect diversity in landraces or cultivated barley in addition to corresponding randomly chosen subsets. All SNP sets except the landrace-optimised subsets underestimated the diversity present in the landrace germplasm, and all subsets of SNP gave similar estimates for cultivar germplasm. All marker subsets gave qualitatively similar estimates of the population structure in both germplasm sets, but the 96 SNP sets showed much lower data resolution values than the larger SNP sets. From these data we deduce that pre-selecting markers for their diversity in a germplasm set is very worthwhile in terms of the quality of data obtained. Second, we suggest that a properly chosen 384 SNP subset gives a good combination of power and economy for germplasm characterization, whereas the rather modest gain from using 1536 SNPs does not justify the increased cost and 96 markers give unacceptably low performance. Lastly, we propose a specific 384 SNP subset as a standard genotyping tool for middle-eastern landrace barley.     

Establishment of a Core Collection of Traditional Cuban <Emphasis Type="Italic">Theobroma cacao</Emphasis> Plants for Conservation and Utilization Purposes

Presently, Theobroma cacao L. (cacao) in Cuba is mainly cultivated in the eastern region where plantations comprise a mixture of clonal varieties, hybrids, progeny of Trinidad Selected Hybrids, and traditional—also known as ancient—cacao. The ancient genetic resources, probably the plants most closely related to the original introductions, are endangered by their progressive replacement by modern clones. To promote the conservation and utilization of these genetic resources, a representative sample of 537 traditional Cuban cacao plants was used to develop a core collection. Core collections based on 15 simple sequence repeat (SSR) markers were generated using five different sampling algorithms: random sampling, simulated annealing, stepwise clustering with random sampling, the M strategy, and maximum genetic diversity. The five core collections were designed to capture 95 % of the SSR alleles in the complete collection. The genetic, morphological, and geographical diversity of each core collection was compared with that of the entire collection. The entire collection contained 139 alleles, including 104 rare ones, with the 95 % allelic coverage threshold achieved with 133 alleles. The core collection generated by the maximum genetic diversity algorithm had the lowest number of accessions (185), the highest mean genetic distance (0.486), the lowest morphological character redundancy, and the highest diversity as assessed by the mean Shannon-Weaver diversity index (0.757). This core collection can thus serve as the basis of future improvement programs based on local genetic resources.     

Comparison of different methods to construct a core germplasm collection in woody perennial species with simple sequence repeat markers. A case study in cherimoya (Annona cherimola, Annonaceae), an underutilised subtropical fruit tree species

Although molecular markers are becoming the tool of choice to develop core collections in plants, the examples of their use in woody perennial species are very scarce. In this work, we used simple sequence repeat (SSR) marker data to develop a core collection in an underutilised subtropical fruit tree species, cherimoya ( Annona cherimola , Annonaceae), from an initial collection of 279 genotypes from different countries. We compared six alternative allocation methods to construct the core collection, four not based upon the similarity dendrogram [random sampling, maximisation strategy (M strategy) and simulated annealing algorithm maximising both genetic diversity and number of SSR alleles] and two based on dendrogram data (logarithmic strategy and stepwise clustering). The diversity maintained in each subset was compared with that present in the entire collection. The results obtained indicate that the use of SSRs together with the M strategy is the most efficient method to develop a core collection in cherimoya. In the best subset, with 40 accessions, all the SSR alleles present in the whole collection were recovered and no significant differences in frequency distribution of alleles for any of the loci studied or in variability parameters ( H _O, H _E) were recorded between the core and the whole collection.     

Maintenance of sorghum (sorghum bicolor, poaceae) landrace diversity by farmers’ selection in Ethiopia

We quantitatively examined the relationships between Sorghum landrace diversity at the field level and environmental factors and farmers’ selection practices in north Shewa and south Welo regions of Ethiopia. Surveys were conducted on 260 randomly selected farmers’ fields. The altitude and size of each field were recorded. Sorghum plants at 5 m intervals along transect lines spaced 10 m apart over each field were identified by the farmers and the owner of each field was asked why she/he decided to grow each plant. Soil samples were collected from all of the fields and analyzed for pH, organic content, and sand, silt and clay content. Simple and polynomial regressions and multiple regression analyses showed that Sorghum landrace diversity at the field level had significant relationships with the number of selection criteria used by the farmers, field altitude, field size, pH and clay content. As the number of selection criteria increased, landrace diversity in the fields increased. This relationship was not a result of the correlation between selection criteria and the environmental factors, because it was significant after statistically correcting for the effects of the environmental variables. This study quantitatively confirms the role of traditional farmers in the maintenance of sorghum landrace diversity in north Shewa and south Welo regions of Ethiopia.     

Optimal sampling strategy and core collection size of Andean tetraploid potato based on isozyme data - a simulation study

Selection of an appropriate sampling strategy is an important prerequisite to establish core collections of appropriate size in order to adequately represent the genetic spectrum and maximally capture the genetic diversity in available crop collections. We developed a simulation approach to identify an optimal sampling strategy and core-collection size, using isozyme data from a CIP germplasm collection on an Andean tetraploid potato. Five sampling strategies, constant (C), proportional (P), logarithmic (L), square-root (S) and random (R), were tested on isozyme data from 9,396 Andean tetraploid potato accessions characterized for nine isozyme loci having a total of 38 alleles. The 9,396 accessions, though comprising 2,379 morphologically distinct accessions, were found to represent 1,910 genetically distinct groups of accessions for the nine isozyme loci using a sort-and-duplicate-search algorithm. From each group, one accession was randomly selected to form a genetically refined entire collection (GREC) of size 1,910. The GREC was used to test the five sampling strategies. To assess the behavior of the results in repeated sampling, k = 1,500 and 5,000 independent random samples (without replacement) of admissible sizes n = 50(50)1,000 for each strategy were drawn from GREC. Allele frequencies (AF) for the 38 alleles and locus heterozygosity (LH) for the nine loci were estimated for each sample. The goodness of fit of samples AF and LH with those from GREC was tested using the L² test. A core collection of size n = 600, selected using either the P or the R sampling strategy, was found adequately to represent the GREC for both AF and LH. As similar results were obtained at k = 1,500 and 5,000, it seems adequate to draw 1,500 independent random samples of different sizes to test the behavior of different sampling strategies in order to identify an appropriate sampling approach, as well as to determine an optimal core collection size.     

Local genetic diversity of sorghum in a village in northern Cameroon: structure and dynamics of landraces

We present the first study of patterns of genetic diversity of sorghum landraces at the local scale. Understanding landrace diversity aids in deciphering evolutionary forces under domestication, and has applications in the conservation of genetic resources and their use in breeding programs. Duupa farmers in a village in Northern Cameroon distinguished 59 named sorghum taxa, representing 46 landraces. In each field, seeds are sown as a mixture of landraces (mean of 12 landraces per field), giving the potential for extensive gene flow. What level of genetic diversity underlies the great morphological diversity observed among landraces? Given the potential for gene flow, how well defined genetically is each landrace? To answer these questions, we recorded spatial patterns of planting and farmers’ perceptions of landraces, and characterized 21 landraces using SSR markers. Analysis using distance and clustering methods grouped the 21 landraces studied into four clusters. These clusters correspond to functionally and ecologically distinct groups of landraces. Within-landrace genetic variation accounted for 30% of total variation. The average F _is over landraces was 0.68, suggesting high inbreeding within landraces. Differentiation among landraces was substantial and significant (F _st = 0.36). Historical factors, variation in breeding systems, and farmers’ practices all affected patterns of genetic variation. Farmers’ practices are key to the maintenance, despite gene flow, of landraces with different combinations of agronomically and ecologically pertinent traits. They must be taken into account in strategies of conservation and use of genetic resources.     

Methods of developing a core collection of annual Medicago species

A core collection is a subset of a large germplasm collection that contains accessions chosen to represent the genetic variability of the germplasm collection. The purpose of the core collection is to improve management and use of a germplasm collection. Core collections are usually assembled by grouping accessions and selecting from within these groups. The objective of this study was to compare 11 methods of assembling a core collection of the U.S. National collection of annual Medicago species. These methods differed in their use of passport and evaluation data as well as their selection strategy. Another objective was to compare core collections with sample sizes of 5%, 10% and 17% of the germplasm collection. Core collections assembled with evaluation data and cluster analysis better represented the germplasm collection than core collections assembled based solely on passport data and random selection of accessions, The Relative Diversity and the logarithm methods generated better core collections than the proportional method. The 5% and 10% sample size core collection were judged insufficient to represent the germplasm collection.     

基于AFLP分子标记的桂花品种核心种质的构建

用100个桂花品种荧光AFLP分子标记信息构建核心种质。利用获得的指纹信息,运用UPGMA聚类取样法,采用Kimura 2-parameter遗传距离,多次聚类随机抽样。结果表明:(1)8对引物共获得514条带,平均每对引物获得64条带。(2)从100个桂花品种中筛选了30个样本的核心种质。(3)比较核心种质和全部种质的Shan-non-Wiener指数(H′)和Simpson指数(D),t检验值均说明核心种质的遗传多样性指数与全部种质遗传多样性没有明显差异,表明所构建的核心样品能够很好地保留原始100个桂花品种的遗传多样性。     

利用ISSR分子标记构建濒危植物水青树核心种质

基于全国26个居群的174份水青树Tetracentron sinense种质,利用14条ISSR引物,采用UPGMA聚类法进行多次逐步聚类,利用三种取样策略(随机取样策略、位点优先取样策略和偏离度取样策略)构建初始核心种质,并将各遗传多样性指标进行比较分析,从而确定构建水青树核心种质的最佳方法。利用14条ISSR引物扩增出180个条带,其观测等位基因数(Na)、有效等位基因数(Ne)、Nei’s基因多样性指数(H)与Shannon信息指数(I)分别为1.6944、1.3912、0.2321和0.3509,说明水青树具有较低的遗传多样性。采用位点优先取样策略构建的核心种质各遗传多样性指数均大于随机取样策略和偏离度取样策略。利用位点优先取样策略构建了78 份核心种质资源,保留了44.83%原种质样品,遗传多样性指数Na、Ne、H和I的保留率分别为98.69%、100.01%、99.27%和98.67%,比较完好地保留了原种质的遗传多样性和遗传变异。因此,位点优先取样策略是构建水青树核心种质的最佳取样策略。     

Assessment of genetic diversity within and among germplasm accessions in cultivated sorghum using microsatellite markers

Microsatellite markers are increasingly being used in crop plants to discriminate among genotypes and as tools in marker-assisted selection. Here we evaluated the use of microsatellite markers to quantify the genetic diversity within as well as among accessions sampled from the world germplasm collection of sorghum. Considerable variation was found at the five microsatellite loci analysed, with an average number of alleles per locus equal to 2.4 within accessions and 19.2 in the overall sample of 25 accessions. The collection of sorghum appeared highly structured genetically with about 70% of the total genetic diversity occurring among accessions. However, differentiation among morphologically defined races of sorghum, or among geographic origins, accounted for less than 15% of the total genetic diversity. Our results are in global agreement with those obtained previously with allozyme markers. We were also able to show that microsatellite data are useful in identifying individual accessions with a high relative contribution to the overall allelic diversity of the collection. Received: 10 August 1999 / Accepted: 27 August 1999     

中国普通小麦初选核心种质的产生

对中国普通小麦种质资源构建了初选核心种质。地方品种和选育品种分别构建。按栽培区(地理生态区)分组。地方品种按亚区分为28组,选育品种按大区分为10组。各组内在21个表型性状聚类的基础上,按平方根法取样,并依遗传多样性指数与遗传丰富度加以调整。提出在生产上或育种中起过重要作用的品种为必选材料。初步选定的材料经种植核对,淘汰错杂后,产生初选核心种质。地方品种全部供试材料11694份,初选核心种质3283份,取样比例为28．18％。选育品种全部11441份,初选核心种质1684份,取样比例为14．9％。计划经分子标记分析,最后核心种质的比例占全部种质的10％左右。根据全部材料21个性状遗传多样性指数测验,初选核心种质,除芒和壳两性状外,与全部种质的遗传差异均未达到显水平。讨论了初选核心种质的构建方法。指出陕南部西山地和汾渭谷地是中国小麦地方品种遗传变异多样性的富集地。育成品种多样性程度以西南冬麦区和黄淮冬麦区为最高。     

An SSR‐based approach incorporating a novel algorithm for identification of rare maize genotypes facilitates criteria for landrace conservation in Mexico

As maize was domesticated in Mexico around 9,000 years ago, local farmers have selected and maintained seed stocks with particular traits and adapted to local conditions. In the present day, many of these landraces are still cultivated; however, increased urbanization and migration from rural areas implies a risk that this invaluable maize germplasm may be lost. In order to implement an efficient mechanism of conservation in situ, the diversity of these landrace populations must be estimated. Development of a method to select the minimum number of samples that would include the maximum number of alleles and identify germplasm harboring rare combinations of particular alleles will also safeguard the efficient ex‐situ conservation of this germplasm. To reach this goal, a strategy based on SSR analysis and a novel algorithm to define a minimum collection and rare genotypes using landrace populations from Puebla State, Mexico, was developed as a “proof of concept” for methodology that could be extended to all maize landrace populations in Mexico and eventually to other native crops. The SSR‐based strategy using bulked DNA samples allows rapid processing of large numbers of samples and can be set up in most laboratories equipped for basic molecular biology. Therefore, continuous monitoring of landrace populations locally could easily be carried out. This methodology can now be applied to support incentives for small farmers for the in situ conservation of these traditional cultivars.     

基于农艺性状指标的山西高粱地方品种核心种质构建

利用1285份山西省高粱地方品种18个农艺性状的历史数据,通过比较不同取样方法、取样比例和聚类方法组合的构建方法,确定了"多次聚类偏离度取样法+15％取样比例+欧氏距离+最长距离法"为山西省高粱地方核心种质构建的方法.192份初选核心种质和所有样本的均值差异百分率、方差差异百分率、极差符合率和变异系数变化率分别为0、8...     

A mini core subset for capturing diversity and promoting utilization of chickpea genetic resources in crop improvement

A core collection is a chosen subset of large germplasm collection that generally contains about 10% of the total accessions and represents the genetic variability of entire germplasm collection. The purpose of a core collection is to improve the use of genetic resources in crop improvement programs. In many crops the number of accessions contained in the genebank are several thousands, and a core subset consisting of 10% of total accessions would be an unwieldy proposition. In this article we have suggested a two-stage strategy to select a chickpea mini core subset consisting of only about 1% of the entire collection held in trust at ICRISAT’s genebank (16,991 accessions). This mini core subset still represents the diversity of the entire core collection. The first stage involves developing a representative core subset (about 10%) from the entire collection using all the available information on origin, geographical distribution, and characterization and evaluation data of accessions. The second stage involves evaluation of the core subset for various morphological, agronomic, and quality traits, and selecting a further subset of about 10% accessions from the core subset. At both stages standard clustering procedure was used to separate groups of similar accessions. A mini core subset consisting 211 accessions from 1,956 core subset accessions, using data on 22 morphological and agronomic traits, was selected. Newman- Keuls’ test for means, Levene’s test for variances, the chi-square test and Wilcoxon’s rank-sum non-parametric test for frequency distribution analysis for different traits indicated that the variation available in the core collection has been preserved in the mini core subset. The most important phenotypic correlations which may be under the control of coadapted gene complexes, were also preserved in the mini core. This mini core subset, due to its drastically reduced size, will prove to be a point of entry to proper exploitation of chickpea genetic resources. Received: 20 August 2000 / Accepted: 25 September 2000     

Genetic characterization of a core set of a tropical maize race Tuxpeño for further use in maize improvement

The tropical maize race Tuxpe?o is a well-known race of Mexican dent germplasm which has greatly contributed to the development of tropical and subtropical maize gene pools. In order to investigate how it could be exploited in future maize improvement, a panel of maize germplasm accessions was assembled and characterized using genome-wide Single Nucleotide Polymorphism (SNP) markers. This panel included 321 core accessions of Tuxpe?o race from the International Maize and Wheat Improvement Center (CIMMYT) germplasm bank collection, 94 CIMMYT maize lines (CMLs) and 54 U.S. Germplasm Enhancement of Maize (GEM) lines. The panel also included other diverse sources of reference germplasm: 14 U.S. maize landrace accessions, 4 temperate inbred lines from the U.S. and China, and 11 CIMMYT populations (a total of 498 entries with 795 plants). Clustering analyses (CA) based on Modified Rogers Distance (MRD) clearly partitioned all 498 entries into their corresponding groups. No sub clusters were observed within the Tuxpe?o core set. Various breeding strategies for using the Tuxpe?o core set, based on grouping of the studied germplasm and genetic distance among them, were discussed. In order to facilitate sampling diversity within the Tuxpe?o core, a minicore subset of 64 Tuxpe?o accessions (20% of its usual size) representing the diversity of the core set was developed, using an approach combining phenotypic and molecular data. Untapped diversity represents further use of the Tuxpe?o landrace for maize improvement through the core and/or minicore subset available to the maize community.