Use of SAMPL for a study of DNA polymorphism,genetic diversity and possible gene tagging in bread wheat

Selective Amplification of Microsatellite Polymorphic Loci (SAMPL) technology was used in bread wheat for the first time for a study of genetic diversity, genotype identification and gene tagging. The diversity studies involved 55 wheat genotypes and two SAMPL primer pairs (SAMPL-6 and SAMPL-7, each with a M-CAG primer), which together gave 43 polymorphic bands out of a total of 87 SAMPL bands. The average polymorphic information content (PIC) of SAMPL primers was 0.221 and that of SAMPL markers was 0.264. The marker index of SAMPL markers was 9.61. The genetic similarity (GS) coefficients for 1,485 pairs of genotypes ranged from 0.35 to 0.96 with an average of 0.65. A dendrogram was prepared on the basis of a similarity matrix using the UPGMA algorithm, which corresponded well with the results of principal component analysis (PCA). From a total of 55 genotypes, 54 could be distinguished using the SAMPL banding patterns of both primers. For gene tagging, 568 bands from a total of 1,185 SAMPL bands detected polymorphism between each of the three pairs of parents differing for grain protein content (GPC), pre-harvest sprouting tolerance (PHST) and grain weight (GW). An association of six bands with GPC, of seven bands with PHST and four bands with GW was observed using bulked segregant analysis (BSA). Received: 5 April 2001 / Accepted: 17 May 2001     

Linkage mapping in apomictic and sexual Kentucky bluegrass ( Poa pratensis L.) genotypes using a two way pseudo-testcross strategy based on AFLP and SAMPL markers

The high versatility of the mode of reproduction and the retention of a pollen recognition system are the factors responsible for the extreme complexity of the genome in Poa pratensis L. Two genetic maps, one of an apomictic and one of a sexual genotype, were constructed using a two-way pseudo-testcross strategy and multiplex PCR-based molecular markers (AFLP and SAMPL). Due to the high ploidy level and the uncertainty of chromosome pairing-behavior at meiosis, only parent-specific single-dose markers (SDMs) that segregated 1:1 in an F₁ mapping population (161 out of 299 SAMPLs, and 70 out of 275 AFLPs) were used for linkage analysis. A total of 41 paternal (33 SAMPLs and 8 AFLPs) and 47 maternal (33 SAMPLs and 14 AFLPs) SDMs, tested to be linked in coupling phase, were mapped to 7+7 linkage groups covering 367 and 338.4 cM, respectively. The comparison between the two marker systems revealed that SAMPL markers were statistically more efficient than AFLP ones in detecting parent-specific SDMs (75% vs 32.4%). There were no significant differences in the percentages of distorted marker alleles detected by the two marker systems (27.8% of SAMPLs vs 21.3% of AFLPs). The pairwise comparison of co-segregational groups for linkage detection between marker loci suggested that at least some of the P. pratensis chromosomes pair preferentially at meiosis-I. Received: 31 August 2000 / Accepted: 12 January 2001     

Development of SCAR Markers for Germplasm Characterisation in Olive Tree (Olea europea L.)

A set of 14 SCAR markers were developed starting from RAPD, AFLP and SAMPL analysis of several olive germplasm accessions. Eight RAPD, two AFLP and four SAMPL fragments were converted into dominant and codominant SCARs by cloning and sequencing the selected fragments. The markers obtained were evaluated on forty different olive cultivars from different Italian production areas (mainly from Liguria). The combined use of these SCARs made possible to univocally identify 26 cultivars while the remaining 14 will require the development of further markers since most of them are placed in a main group containing six genetically similar cultivars (among which Frantoio and Taggiasca) and four minor groups containing two cultivars each. A total of 31 different haplotypes were identified and the analysis of several individual plants indicated no intra-cultivar variability. Considering the SCAR polymorphism two alleles were scored for each markers with the only exception of markers IGPS3 and IGPS4 showing 4 alleles with 7 recognised groups and 5 alleles with 4 groups, respectively. Though less polymorphic in comparison with other markers like SSRs, the developed SCARs proved useful in genotype identification. In addition, they could potentially be used for breeding applications and forensic analysis. An erratum to this article is available at .     

Linkage relationships among molecular markers and storage root traits of carrot (Daucus carota L. ssp. sativus)

 A 109-point linkage map consisting of three phenotypic loci (P ₁, Y ₂, and Rs), six restriction fragment length polymorphisms (RFLPs), two random amplified polymorphic DNAs (RAPDs), 96 amplified fragment length polymorphisms (AFLPs), and two selective amplification of microsatellite polymorphic loci (SAMPL) was constructed for carrot (Daucus carota L. ssp. sativus; 2n=2x=18). The incidence of polymorphism was 36% for RFLP probes, 20% for RAPD primers, and 42% for AFLP primers. The overall incidence of disturbed segregation was 18%. Linkage relationships at a LOD score of 4.0 and θ=0.25 indicated 11 linkage groups. The total map length was 534.4 cM and the map was clearly unsaturated with markers spaced at 4.9 cM. AFLP P6B15 was 1.7 cM from P ₁, AFLP P1B34 was 2.2 cM from Y ₂, and AFLP P3B30XA was 8.1 cM from Rs. Received: 2 September 1998 / Accepted: 28 November 1998     

A first linkage map of Cichorium intybus L. using a one-way pseudo-testcross and PCR-derived markers

We have used a one-way pseudo-testcross mapping strategy in combination with different types of PCR-based markers (RAPD, AFLP, SAMPL) to construct a first linkage map for variegated chicory (Cichorium intybus L. var. silvestre Biskoff, n=9), a self-incompatible vegetable species. The success of such a strategy depends on the presence of sufficiently high levels of heterozygosity in the individual plant which is being mapped and on the informativeness of the marker system that is used. A total of 371 markers, comprising 16 RAPDs, 72 SAMPLs and 283 AFLPs, were scored in 46 F₁ individuals obtained from an interspecific cross between a C. intybus outbred individual and a C. endivia inbred line. Grouping of the markers at a LOD score of 4.0 resulted in 13 linkage groups covering 1330 cM. A framework map covering 1201.4 cM was assembled by using all markers that could be ordered with a LOD greater than 2.0. We estimate the total genome size of chicory to be ca. 1405 cM, thus considerably smaller than that estimated for lettuce (1950 cM). The usefulness of the different marker systems that were applied is analysed in terms of level of heterozygosity and marker index, i.e. number of different genetic loci that may be simultaneously analysed per experiment. Out of the 371 markers, 50 of them showed segregation distortion which is discussed in terms of the hybrid origin of the variegated chicory.     

Large scale development of selectively amplified microsatellite polymorphic loci (SAMPL) markers in spruce (Picea)

The spruce (Picea) species are ecologically and economically important in Canada. Highly informative markers with high multiplex ratios are needed to assist spruce genomics, genetics, and breeding programs. Selectively amplified microsatellite polymorphic loci (SAMPL) markers are highly suitable for these programs. We have developed, optimized, and characterized a set of 10 new SAMPL primers in combination with 16 MseI primers and resolved a large number of polymorphic SAMPL markers in spruce. The SAMPL primers were designed from the compound microsatellite repeats found in Norway spruce (Picea abies) and white spruce (Picea glauca). A total of 6313 polymorphic SAMPL makers were produced by 160 SAMPL–MseI primers combinations in eight progeny of a spruce mapping population.     

Correlations between Genetic,Morphological, and Chemical Diversities in a Germplasm Collection of the Medicinal Plant Origanum vulgare L.

In total, 42 accessions of Origanum vulgare L., mostly originating from Europe, were evaluated, to detect molecular, quantitative morphological, and chemotype polymorphisms and to discover possible correlations between them. Twelve traits related to morphological characteristics were measured. The components in the essential oils were identified by GC/MS analysis, and the oil contents of 18 major compounds were determined. A total of 477 molecular polymorphisms including 214 AFLP (amplified fragment length polymorphism) and 263 SAMPL (selectively amplified microsatellite polymorphic loci) were used for genotyping. Euclidean distances of morphological and chemotypic data and genetic distances (1 – Dice's similarity) of molecular markers were compared by applying Mantel tests to ascertain the congruencies between them. A relatively high correlation between chemotypic patterns and genetic markers was identified, while a lower correlation was found between the morphological and genetic matrices. Pairwise analyses of correlation among all traits showed that the stem diameter was correlated to the essential‐oil yield and the carvacrol content. Cluster analysis, population inference, and principal component analysis revealed a broad genetic and chemical variation among the accessions. The knowledge of these diversities, found in this study, will allow a plant improvement of Origanum vulgare related to pharmaceutical and spice uses.     

Towards second-generation STS (sequence-tagged sites) linkage maps in conifers: a genetic map of Norway spruce ( Picea abies K.)

Genetic linkage maps have been produced for a wide range of organisms during the last decade, thanks to the increasing availability of molecular markers. The use of microsatellites (or Simple Sequence Repeats, SSRs) as genetic markers has led to the construction of “second-generation” genetic maps for humans, mouse and other organisms of major importance. We constructed a second-generation single-tree genetic linkage map of Norway spruce (Picea abies K.) using a panel of 72 haploid megagametophytes with a total of 447 segregating bands [366 Amplified Fragment Length Polymorphisms (AFLPs), 20 Selective Amplification of Microsatellite Polymorphic Loci (SAMPLs) and 61 SSRs, each single band being treated initially as a dominant marker]. Four hundred and thirteen markers were mapped in 29 linkage groups (including triplets and doublets) covering a genetic length of 2198.3 cM, which represents 77.4% of the estimated genome length of Picea abies (approximately 2839 cM). The map is still far from coalescing into the expected 12 chromosomal linkage groups of Norway spruce (2n = 2x = 24). A possible explanation for this comes from the observed non-random distribution of markers in the framework map. Thirty-eight SSR marker loci could be mapped onto 19 linkage groups. This set of highly informative Sequence Tagged Sites (STSs) can be used in many aspects of genetic analysis of forest trees, such as marker-assisted selection, QTL mapping, positional cloning, gene flow analysis, mating system analysis and genetic diversity studies. Received: 5 November 1997 / Accepted: 16 March 1998     

SAMPL: A technique for somaclonal variation fingerprinting inMusa

SAMPL primers designed for genomic profiling in chickpea (Cicer arietinum L.) were tested for their applicability to fingerprinting of DNA of banana cultivars and soma-clonal variants. Most of the chickpea primers allowed amplification of genomic DNA of banana and detection of sequence polymorphisms within theMusa acuminata genome (A). Our results demonstrate that the highly resolving SAMPL technique is useful in banana genomics, especially for the distinction and characterization of commercially important cultivars and promising somaclonal variants containing the A genome.