Evaluation of a DNA pooled-sampling strategy for estimating the RFLP diversity of maize populations

Molecular characterization by means of RFLPs of large sets of populations is presently limited by experimental costs. In order to reduce costs, we have evaluated a method based on the RFLP analysis of balanced bulks of DNA from several individuals. The precision of this approach for estimating allele frequencies within each population was investigated using (i) DNA extracted from controlled bulks of leaf tissues of maize inbred lines, (ii) data obtained from individual analyses of 10 maize populations. Both approaches showed that allele frequencies can generally be estimated with a high precision (coefficients of determination up to 0.99 for some probe-enzyme combinations assayed), relative to the variation in allele frequencies observed among maize populations. Although further efforts are needed to define a set of probe-enzyme combinations that can be routinely image processed, these results, and preliminary results from a 65 maize populations project, suggest that this approach could provide highly informative data for large sets of populations, and at relatively low costs.     

EST–SSR markers for asparagus genetic diversity evaluation and cultivar identification

Simple sequence repeat (SSR) markers generated from expressed sequence tag (EST) sequences represent useful tools for genotyping and their development is relatively easy because of the public availability of EST databases. We report design and application of EST–SSRs to assess the level of genetic diversity among thirty-five asparagus cultivars and to fingerprint DePaoli, a new variety released by University of California, Riverside. DNA was isolated from bulks of pooled cladophylls coming from five plants of each variety to reduce the number of DNA extractions and PCR reactions. Allele frequencies were estimated from the intensity of the bands in two bulks and two individual plant samples for each variety. Although asparagus varieties derive from a limited germplasm pool, eight EST–SSR loci differentiated all of the analyzed cultivars. Moreover, UPGMA (unweighted pair group method with arithmetic mean) and neighbor-joining trees, as well as principal components analysis separated the cultivars into clusters corresponding to the geographical areas where they originated.     

Use of SSRs for establishing heterotic groups in subtropical maize

Heterotic groups and patterns are of fundamental importance in hybrid breeding. The objectives of our research were to: (1) investigate the relationship of simple sequence repeats (SSR) based genetic distances between populations and panmictic midparent heterosis (PMPH) in a broad range of CIMMYT maize germplasm, (2) evaluate the usefulness of SSR markers for defining heterotic groups and patterns in subtropical germplasm, and (3) examine applications of SSR markers for broadening heterotic groups by systematic introgression of other germplasm. Published data of two diallels and one factorial evaluated for grain yield were re-analyzed to calculate the PMPH in population hybrids. Additionally, 20 pools and populations widely used in CIMMYT's breeding program were assayed with 83 SSR markers covering the entire maize genome. Correlations of squared modified Roger's distance (MRD²) and PMPH were mostly positive and significant, but adaption problems caused deviations in some cases. For intermediate- and early-maturity subtropical germplasm, two heterotic groups could be suggested consisting of a flint and dent composite. We concluded that the relationships between the populations obtained by SSR analyses are in excellent agreement with pedigree information. SSR markers are a valuable complementation to field trials for identifying heterotic groups and can be used to introgress exotic germplasm systematically.Communicated by F. Salamini     

利用分子标记分析遗传多样性时的玉米群体取样策略研究

利用分子标记技术对玉米种质资源进行遗传多样性分析对种质资源的保存和利用具有重要的指导意义。但是,在对地方品种和育种群体这些开放授粉群体进行大规模遗传多样性分析时,取样方法将会严重影响到研究结果和工作效率。本研究用2个育种群体和3个地方品种为试材,利用微卫星(SSR)标记对每个群体100个个体及其组成的不同随机混合样品进行了分子检测。结果表明,不同群体的群体内遗传变异大小存在差异;相同数目的个体随机混合的不同样品间的检测结果基本相同;不同数目的个体混合的样品间存在一定程度的差异,并且与材料本身的遗传变异大小有一定关系。考虑到结果的科学性和工作的可行性,建议在利用分子标记(如SSR)进行地方品种和育种群体的遗传多样性评估时,随机选取30个个体组成混合样(或用15个个体组成2个混合样)来代表1个地方品种或育种群体进行分子鉴定。     

Identification of RAPD markers linked to a fertility restorer gene for theOgura radish cytoplasmic male sterility of rapeseed (Brassica napus L.)

Bulked segregant analysis was employed to identify random amplified polymorphic DNA (RAPD) markers linked to the restorer gene (Rfo) used in theOgura radish cytoplasmic male sterility of rapeseed. A total of 138 arbitrary 10-mer oligonucleotide primers were screened on the DNA of three pairs of bulks, each bulk corresponding to homozygous restored and male sterile plants of three segregating populations. Six primers produced repeatable polymorphisms between paired bulks. DNA from individual plants of each bulk was then used as a template for amplification with these six primers. DNA polymorphisms generated by four of these primers were found to be completely linked to the restorer gene with the polymorphic DNA fragments being associated either with the fertility restorer allele or with the sterility maintainer allele. Pairwise cross-hybridization demonstrated that the four polymorphic DNA fragments did not share any homology. Southern hybridization of labelled RAPD fragments on digested genomic DNA from the same three pairs of bulks revealed fragments specific to either the male sterile bulks or to the restored bulks and a few fragments common to all bulks, indicating that the amplified sequences are low copy. The four RAPD fragments that were completely linked to the restorer locus have been cloned and sequenced to develop sequence characterized amplified regions (SCARs). This will facilitate the construction of restorer lines used in breeding programs and is the first step towards map-based cloning of the fertility restorer allele.     

Bulk segregant analysis with molecular markers and its use for improving drought resistance in maize

The usual method to locate and compare loci regulating quantitative traits (QTLs) requires a segregating population of plants with each one genotyped with molecular markers. However, plants from such segregating populations can also be grouped according to phenotypic expression of a trait and tested for differences in allele frequency between the population bulks: bulk segregant analysis (BSA). The same probes used for making a genetic map (e.g. isozyme, RFLP, RAPD, etc) can be used for BSA. A molecular marker showing polymorphism between the parents of the population and which is closely-linked to a major QTL regulating a particular trait will mainly co-segregate with that QTL, i.e. segregate according to the phenotype if the QTL has a large effect. Thus, if plants are grouped according to expression of the trait and extreme groups tested with that polymorphic marker, the frequency of the two marker alleles present within each of the two bulks should deviate significantly from the ratio of 1 : 1 expected for most populations. As chromosomal locations of many molecular markers have now been determined in many species, the map location of closely-linked QTLs can therefore be deduced without having to genotype every individual in segregating populations. This has been used successfully with composite populations of maize to locate QTLs associated with yield under severe drought. An inbred line derived from one of the populations selected for higher drought yield has been crossed with a drought-susceptible inbred line to produce a mapping population for QTL analysis of physiological and developmental traits likely to regulate yield under drought. Future work to identify traits having QTLs with flanking markers showing significant allele frequency differences in the GSA studies will indicate those traits likely to be important in determining yield under drought.Key words: Bulk segregant analysis (BSA), drought resistance, genetic maps, maize, molecular markers, Zea mays (L.).      

A comparison of simple sequence repeat and single nucleotide polymorphism marker technologies for the genotypic analysis of maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.)

We report on the comparative utilities of simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers for characterizing maize germplasm in terms of their informativeness, levels of missing data, repeatability and the ability to detect expected alleles in hybrids and DNA pools. Two different SNP chemistries were compared; single-base extension detected by Sequenom MassARRAY, and invasive cleavage detected by Invader chemistry with PCR. A total of 58 maize inbreds and four hybrids were genotyped with 80 SSR markers, 69 Invader SNP markers and 118 MassARRAY SNP markers, with 64 SNP loci being common to the two SNP marker chemistries. Average expected heterozygosity values were 0.62 for SSRs, 0.43 for SNPs (pre-selected for their high level of polymorphism) and 0.63 for the underlying sequence haplotypes. All individual SNP markers within the same set of sequences had an average expected heterozygosity value of 0.26. SNP marker data had more than a fourfold lower level of missing data (2.1-3.1%) compared with SSRs (13.8%). Data repeatability was higher for SNPs (98.1% for MassARRAY SNPs and 99.3% for Invader) than for SSRs (91.7%). Parental alleles were observed in hybrid genotypes in 97.0% of the cases for MassARRAY SNPs, 95.5% for Invader SNPs and 81.9% for SSRs. In pooled samples with mixtures of alleles, SSRs, MassARRAY SNPs and Invader SNPs were equally capable of detecting alleles at mid to high frequencies. However, at low frequencies, alleles were least likely to be detected using Invader SNP markers, and this technology had the highest level of missing data. Collectively, these results showed that SNP technologies can provide increased marker data quality and quantity compared with SSRs. The relative loss in polymorphism compared with SSRs can be compensated by increasing SNP numbers and by using SNP haplotypes. Determining the most appropriate SNP chemistry will be dependent upon matching the technical features of the method within the context of application, particularly in consideration of whether genotypic samples will be pooled or assayed individually.     

PPC: an algorithm for accurate estimation of SNP allele frequencies in small equimolar pools of DNA using data from high density microarrays

Robust estimation of allele frequencies in pools of DNA has the potential to reduce genotyping costs and/or increase the number of individuals contributing to a study where hundreds of thousands of genetic markers need to be genotyped in very large populations sample sets, such as genome wide association studies. In order to make accurate allele frequency estimations from pooled samples a correction for unequal allele representation must be applied. We have developed the polynomial based probe specific correction (PPC) which is a novel correction algorithm for accurate estimation of allele frequencies in data from high-density microarrays. This algorithm was validated through comparison of allele frequencies from a set of 10 individually genotyped DNA's and frequencies estimated from pools of these 10 DNAs using GeneChip 10K Mapping Xba 131 arrays. Our results demonstrate that when using the PPC to correct for allelic biases the accuracy of the allele frequency estimates increases dramatically.     

Estimation of allele frequency in pooled DNA by using PCR–RFLP combined with microchip electrophoresis

Biallelic marker, most commonly single nucleotide polymorphism (SNP), is widely utilized in genetic association analysis, which can be speeded up by estimating allele frequency in pooled DNA instead of individual genotyping. Several methods have shown high accuracy and precision for allele frequency estimation in pools. Here, we explored PCR restriction fragment length polymorphism (PCR–RFLP) combined with microchip electrophoresis as a possible strategy for allele frequency estimation in DNA pools. We have used the commercial available Agilent 2100 microchip electrophoresis analysis system for quantifying the enzymatically digested DNA fragments and the fluorescence intensities to estimate the allele frequencies in the DNA pools. In this study, we have estimated the allele frequencies of five SNPs in a DNA pool composed of 141 previously genotyped health controls and a DNA pool composed of 96 previously genotyped gastric cancer patients with a frequency representation of 10–90% for the variant allele. Our studies show that accurate, quantitative data on allele frequencies, suitable for investigating the association of SNPs with complex disorders, can be estimated from pooled DNA samples by using this assay. This approach, being independent of the number of samples, promises to drastically reduce the labor and cost of genotyping in the initial association analysis.     

Development of multiplex sets of simple sequence repeat DNA markers covering the soybean genome

Multiplexing involves the analysis of several markers in a single gel lane that is based on the allele size range of marker loci. Multiplex SSR marker analysis is conducted with primers that are labeled with one of three dyes. The development of an SSR multiplex system requires estimates of the allele size range of markers to strategize primer labeling and for grouping markers into multiplex sets. A method is presented that describes the development of multiplex sets of SSR markers in soybean (Glycine max (L.) Merr.) by the selective placement of primer sites and by the analysis of diverse germplasm. Primer sites were placed at specific distances from the SSR to adjust the allele size range of marker loci. The analysis of pooled DNA samples comprising diverse soybean genotypes provided robust estimates of the allele size range of marker loci that enabled the development of multiplex sets. Eleven multiplex sets comprising 74 SSR markers distributed across the 20 linkage groups of soybean were developed. Multiplex sets constructed from the analysis of diverse soybean germplasm should have a wide range of genotyping applications. The procedures used in this study were systematic and rapid and should be applicable for multiplex development in any species with SSR marker technology.     

Estimation of allele frequencies in polyploids under certain patterns of inheritance

Allele frequencies have long been studied by biologists interested in evolution and speciation. More recently, with the application of molecular markers in human DNA profiling we have also seen the need for reliable population allele frequency estimates for making probabilistic inferences. There is now interest in applying the same DNA profiling technology to identification of plant varieties. HortResearch maintains a large germplasm of horticultural plant species. It is becoming evident that accurate identification of these accessions through DNA fingerprinting is essential for effective utilisation and maintenance of this germplasm. Microsatellites are the markers of choice for this fingerprinting. However, such markers do not reveal the dosage of alleles in a polyploid. Polyploidy is common amongst horticultural plants. Estimating allele frequencies in a polyploid population is, therefore, complicated because of some marker genotypes being phenotypically indistinguishable. For example, in a tetraploid, with four alleles at a locus showing polysomic inheritance, although 35 genotypes are possible, these will fall into only 15 marker phenotypic classes. Furthermore 'null' individuals are rarely detected in polyploids. Furthermore, some polyploids can be cryptic exhibiting disomy, instead of the polysomic inheritance. We will discuss the implications of these factors and present an EM-type algorithm for estimating allele frequencies of a polyploid population under certain patterns of inheritance. The method will be demonstrated on simulated data. We also discuss the nature of some of the additional problems that may be encountered with estimating allele frequencies in polyploids for which other solutions still need to be developed.     

Development and mapping of SSR markers for maize

Microsatellite or simple sequence repeat (SSR) markers have wide applicability for genetic analysis in crop plant improvement strategies. The objectives of this project were to isolate, characterize, and map a comprehensive set of SSR markers for maize (Zea mays L.). We developed 1051 novel SSR markers for maize from microsatellite-enriched libraries and by identification of microsatellite-containing sequences in public and private databases. Three mapping populations were used to derive map positions for 978 of these markers. The main mapping population was the intermated B73 × Mo17 (IBM) population. In mapping this intermated recombinant inbred line population, we have contributed to development of a new high-resolution map resource for maize. The primer sequences, original sequence sources, data on polymorphisms across 11 inbred lines, and map positions have been integrated with information on other public SSR markers and released through MaizeDB at URL:www.agron.missouri.edu. The maize research community now has the most detailed and comprehensive SSR marker set of any plant species.     

Molecular tagging of gene conferring leaf blight resistance using microsatellites in sorghum [Sorghum bicolor (L.) Moench

Resistance to leaf blight in sorghum [Sorghum bicolor (L.) Moench] accession G-118 was found to segregate as a single dominant trait in a cross to susceptible cultivar, HC-136. Molecular marker(s) linked to the locus for disease resistance was identified using simple sequence repeat (SSR) markers coupled with bulk segregant analysis. Genomic DNA from the parental cultivars and bulks were screened by PCR amplification with 50 simple sequence repeat primer pairs. Out of these, 38 SSR primers produced polymorphism between parents. After screening of these 38 SSRs with resistant and susceptible bulk, one SSR primer, Xtxp 309 produced a unique band of approximately 700 bp only in resistant parent and resistant bulk and a unique band of 450 bp only in susceptible parent and susceptible bulk. Upon screening with individual resistant and susceptible recombinant inbred lines (RILs), marker Xtxp 309 produced amplification in 23 of the 26 resistant RILs and no amplification was produced in any of the 25 susceptible RILs. The same marker Xtxp 309 produced amplification in 21 of the susceptible RILs and 3 of the resistant RILs of 450 bp band. This was found to be located at a distance of 3.12 cM away from the locus governing resistance to leaf blight which was considered to be closely linked and 7.95 cM away from the locus governing susceptibility to leaf blight. This marker may prove useful in MAS for gene introgression, plant genetic diagnostics and gene pyramiding for resistance via genetic transformation for disease resistance in plants.     

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.     

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.     

A Multiplex Microsatellite Marker Kit for Diversity Assessment of Large Cassava (<Emphasis Type="Italic">Manihot esculenta</Emphasis> Crantz) Germplasm Collections

Current methods for molecular fingerprinting of cassava (Manihot esculenta Crantz) have limited throughput or are costly, thus preventing the characterization of large germplasm collections such as those held by the International Agricultural Research Centers or National Research Institutions, which comprise hundreds to thousands of accessions. Here, we report the development of a fluorescence-based multiplex simple sequence repeat (SSR) marker kit that enables accurate and cost-effective cassava fingerprinting. The kit comprises 16 SSR markers assembled into five multiplex panels and was tested on 21 cassava cultivars alongside one accession of Manihot epruinosa, a wild relative. A total of 68 alleles were detected with, on average, 4.25 alleles per locus and a polymorphism information content of 0.53. The marker kit reported here is comparable to previously published amplified fragment length polymorphism and SSR marker systems in terms of discriminating power and informativeness while offering significant advantages in speed and cost of marker analysis. Previous molecular genetic diversity studies have suggested that cassava germplasm collections contain duplicate entries based on the occurrence of identical genetic profiles. Using the newly developed microsatellite kit, three out of six putative duplicate accessions could be readily differentiated, showing that these are distinct genotypes. The relevance of these findings with respect to the characterization and management of large cassava germplasm collections is discussed.     

Genetic Characterization of a Core Set of a Tropical Maize Race Tuxpe?o for Further Use in Maize Improvement

The tropical maize race Tuxpeño 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 Tuxpeño 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 Tuxpeño core set. Various breeding strategies for using the Tuxpeño core set, based on grouping of the studied germplasm and genetic distance among them, were discussed. In order to facilitate sampling diversity within the Tuxpeño core, a minicore subset of 64 Tuxpeño 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 Tuxpeño landrace for maize improvement through the core and/or minicore subset available to the maize community.     

Use of a novel outbred by inbred F1 cross to detect genetic markers for growth

A unique outbred by inbred F1 resource population was established. The population structure facilitated the unique opportunity of examining gene by genetic background interaction through crossing two modern broiler sires with dams from two unrelated inbred lines, with no selection for growth rate, to produce about 600 F1 chicks. Pools of DNA were generated from the phenotypic extremes (20% high and low) for 8-week body weight for each of the four combinations of sire and dam line. For one sire family, pools were also separately generated for each sex. The pools were genoyped with 25 informative (segregating) microsatellites. This unique F1 cross between outbred and inbred populations allowed use of the inbred alleles as an 'internal control' for polymerase chain reaction amplification quality in DNA pools. Ten microsatellites showed marked differences (P < 0.05) in allele frequencies between high and low pools, suggesting an association between marker and quantitative trait loci (QTL). These differences were verified using selective genotyping. For many markers, differences in allele frequencies between the high and the low pools, or marker effect, varied between the two dam lines and the two sexes, suggesting an interaction between some genes and the genetic background as represented by different dam lines or sexes. The suggestive marker-QTL associations identified in this F1 population demonstrate the efficiency of this population design while different QTL effects in different genetic line crosses and sexes highlight the importance of gene by genetic background interaction in QTL detection.     

Informative SSR markers for commercial variety discrimination in watermelon (Citrullus lanatus)

SSR markers were used for variety discrimination and genetic assessment in watermelon varieties. Genetic characterization of 49 watermelon varieties was investigated using 30 SSR markers developed from melon and watermelon. A total of 121 polymorphic amplified fragments were obtained by using 30 SSR markers. The average polymorphism information content (PIC) was 0.502 ranging from 0.223 to 0.800. One hundred twenty one SSR loci were used to calculate Jaccard’s distance coefficients for unweighted pair group method using the arithmetic averages (UPGMA) cluster analysis. A clustering group of varieties, based on the results of SSR analysis, were categorized into 5 major groups corresponding to morphological traits. Inheritance mode of 2 SSR markers was investigated to F₁ plants and F₂ populations of 2 crosses. Parental alleles were transmitted from F1 plants and F2 populations. Therefore, these marker sets may prove to be effectively applicable to genetic assessment of germplasm, genome mapping, and fingerprinting of watermelon varieties.     

Genetic differentiation analysis of African cassava (Manihot esculenta) landraces and elite germplasm using amplified fragment length polymorphism and simple sequence repeat markers

Molecular‐marker‐aided evaluation of germplasm plays an important role in defining the genetic diversity of plant genotypes for genetic and population improvement studies. A collection of African cassava landraces and elite cultivars was analysed for genetic diversity using 20 amplified fragment length polymorphic (AFLP) DNA primer combinations and 50 simple sequence repeat (SSR) markers. Within‐population diversity estimates obtained with both markers were correlated, showing little variation in their fixation index. The amount of within‐population variation was higher for landraces as illustrated by both markers, allowing discrimination among accessions along their geographical origins, with some overlap indicating the pattern of germplasm movement between countries. Elite cultivars were grouped in most cases in agreement with their pedigree and showed a narrow genetic variation. Both SSR and AFLP markers showed some similarity in results for the landraces, although SSR provided better genetic differentiation estimates. Genetic differentiation (Fst) in the landrace population was 0.746 for SSR and 0.656 for AFLP. The molecular variance among cultivars in both populations accounted for up to 83% of the overall variation, while 17% was found within populations. Gene diversity (H_e) estimated within each population varied with an average value of 0.607 for the landraces and 0.594 for the elite lines. Analyses of SSR data using ordination techniques identified additional cluster groups not detected by AFLP and also captured maximum variation within and between both populations. Our results indicate the importance of SSR and AFLP as efficient markers for the analysis of genetic diversity and population structure in cassava. Genetic differentiation analysis of the evaluated populations provides high prospects for identifying diverse parental combinations for the development of segregating populations for genetic studies and the introgression of desirable genes from diverse sources into the existing genetic base.