Random Amplified Microsatellite Markers Research Articles

Population Genetics of Narcissus Species Reveals High Diversity and Multiple Introductions into Kashmir

Kashmir valley is rich in flora and fauna that is mostly fragmented. Among various ornamental plants, Narcissus is considered exotic and poses threat to local biodiversity. Neutral molecular markers have been exploited for providing information on the diversity and population structure of various invasive and native populations. In the present investigation, 40 samples belonging to five different Narcissus species were genotyped using 13 random amplified microsatellite markers. Dendrogram and principal component analysis revealed high genetic diversity without congruence on geographical or species level. Analysis of molecular variance revealed maximum variability within (93.49%) the Narcissus populations. Fixation indices at the population level were low except for N. poeticus and N. tazetta, indicating slight genetic differentiation in them. Pairwise FST comparisons between five Narcissus populations were low to high. The high diversity observed in the present investigation indicates that the Narcissus species have not gone through any bottleneck due to either multiple introductions or the presence of these species from decades. To ascertain the exotic nature of Narcissus species, a thorough investigation into the species grown in the valley and Narcissus species grown in the centre of origin based on neutral markers is needed.

Characterization of lulo (Solanum quitoense Lam.) genetic diversity in the department of Boyaca, Colombia

Lulo ( Solanum quitoense Lam.) is an exotic fruit with major economic importance in the Andean region. It has gained importance in the agricultural sector due to organoleptic and cultivation characteristics. There are not studies on this plant genetic resource in the province of Neira, Boyaca department, Colombia. Given these concerns, the aim of this research was to characterize the genetic diversity with random amplified microsatellite markers (RAM). Twenty-one lulo materials were collected in the Soaquira and Sacaneca, Boyaca-Colombia districts to calculate genetic diversity and similarity indices. In addition, the analysis with the Nei-Li coefficient formed three groups at 0.60 similarity level showing a loose distribution of the materials without a relationship with the geographical site or the presence or absence of thorns. The seven RAM markers generated 346 alleles, ranging between 43 and 48 for CT and TG, respectively. The average estimated heterozygosity values ranged from 0.33 for TG and 0.40 for CGA. The average value for all evaluated lulo materials was 0.36, higher than the reported value in other studies on lulo genetic diversity in Colombia. The average Fst value for the 20 evaluated materials was 0.14, showing a moderate genetic differentiation. Overall, the RAM markers identified genetic variability in the study area, the need for levels of subdivision and hierarchization greater than those considered in this research, which would have allowed a better use of the plant genetic diversity present in the lulo materials in the province of Neira-Boyaca, Colombia.

Caracterización molecular de cubios (Tropaeolum tuberosum Ruíz y Pavón) en el departamento de Boyacá

El cubio (Tropaeolum tuberosum Ruiz y Pavón) es un tubérculo básico en la dieta de las comunidades andinas. Es un cultivo rústico, con buena adaptación y usos medicinales. Los agricultores de las zonas altas de Boyacá conservan cultivares de tubérculos como cubios, rubas e ibias, que hacen parte de su patrimonio cultural, sin embargo, en los últimos años, este germoplasma está siendo amenazado por la erosión genética causada por factores ambientales y antrópicos. No existen estudios de caracterización molecular de este germoplasma en Boyacá, por lo cual esta investigación se orientó hacia la identificación de la diversidad de once genotipos de cubios, procedentes de los municipios de Ventaquemada y San Pedro de Iguaque, utilizando siete marcadores Microsatélites Amplificados al Azar (RAMs). El análisis de similitud diferenció a los cultivares en dos grandes grupos, de acuerdo con características relacionadas con el tubérculo (color y forma). Los valores de heterocigosidad estimada estuvieron comprendidos entre 0,25 y 0,42 para los cebadores CGA y AG, respectivamente, lo cual es bajo si se compara con los resultados obtenidos en otras especies tuberosas. El valor promedio de heterocigosidad de la población fue de 0,35 y junto con el análisis de varianza molecular y el Fst muestran la existencia de variabilidad genética en los cultivares evaluados, la cual debe tenerse en cuenta en los programas de conservación y mejoramiento genético de los tubérculos andinos.

Caracterización molecular con microsatélites amplificados al azar (RAMs) de Inchi (Caryodendron orinocense K.)

<p><strong><em></em></strong><strong>Título en ingles: Molecular characterization with random amplified microsatellites (RAMs) of Inchi </strong><strong>(<em>Caryodendron orinocense</em> K.)</strong><br /><strong>Resumen: </strong>El Inchi o Cacay (<em>Caryodendron orinocense</em> Karsten) es una de las especies más promisorias de la Amazonía y la Orinoquia colombiana. El principal producto del Cacay son sus almendras, de las que se extrae un aceite con aplicaciones cosméticas, fitoterapéuticas y alimenticias, además presenta un alto contenido de antioxidantes como los Omega 3, 6 y 9 y Vitaminas como la A y E. No existen estudios sobre la caracterización molecular de este recurso fitogenético, por lo cual el objetivo de esta investigación fue caracterizar la diversidad genética usando marcadores Microsatélites Amplificados al Azar (RAMs). El análisis de similitud al 0.50 formó cuatro grupos de acuerdo al sitio geográfico, siendo los materiales procedentes de Putumayo, Cacayal 19, Pauna y Castilla los de menor similitud. Los valores de heterocigosidad estimada fueron de 0.16 y 0.28 para los cebadores CGA y GT, respectivamente. El porcentaje de loci polimórfico varió entre 55% para el cebador CGA y el 90% para el GT. El valor de Fst promedio para los 27 materiales estudiados fue de 0.35, mostrando que la dinámica espacio-temporal de los materiales de <em>Caryodendron </em>tienden hacia una diferenciación genética, propio de sus procesos evolutivos e incidencia de la domesticación.<strong></strong><strong></strong></p><p><strong>Palabras clave: </strong>cacay, marcadores moleculares, diversidad genética, flujo genético, domesticación.<strong></strong></p><p><strong>Abstract: </strong>The Inchi or Cacay (<em>Caryodendron orinocense</em> Karsten) is one of the most promising species of the Amazon and Orinoco Colombian. The main product of Cacay are its almonds, from extracted oil cosmetic, phytotherapeutic and food applications, also has a high content of antioxidants such as Omega 3, 6 and 9 and vitamins like A and E. There are no studies on the molecular characterization of this plant genetic resource; therefore the objective of this research was to characterize the genetic diversity using Random Amplified Microsatellite markers (RAMs). The similarity analysis to 0.50 formed four groups according to geographical location, being materials from Putumayo, Cacayal 19, Pauna and Castilla lowest similarity. Estimated heterozygosity values were 0.16 and 0.28 for the primers CGA and GT, respectively. The percentage of polymorphic loci ranged from 55% for the primer CGA and 90% for the GT. The average Fst value for the 27 materials studied was 0.35, showing the space-temporal dynamics of materials <em>Caryodendron</em><em> </em>tend toward genetic differentiation, due to their own evolutionary processes and domestication incidence.<strong></strong><strong></strong></p><p><strong>Key words:</strong> cacay, molecular markers, genetic diversity, genetic flow, domestication.</p><p><strong>Recibido: </strong>septiembre 15 de 2014<strong> Aprobado: </strong>abril 10 de 2015</p>

English

Eight Random Amplified Microsatellite markers (RAMs) were used to characterize the genetic diversity found in 14 Prunus materials belonging to the deciduous collection of the Pedagogical and Technological University of Colombia. A total of 121 bands were generated: they range from nine for the GT primer to 26 for the ACA primer, and have molecular weights between 100 and 2050 Kb. At a Nei-Li similarity level of 0.75, four groups were formed, according to the characteristics of the fruit. The number of polymorphic loci varied between 8 (GT and AG) and 21 (ACA); the higher levels of heterozygosity were CA (0.43) and CT (0.41). The average value of heterozigosity for the total population was 0.35, much lower than those found in other Prunus species, but higher than other fruit species where RAM markers were used. Therefore, strategies must be deployed for the collections in order to increase genetic variability, such as the introduction of wild or hybrid materials. The RAM technique proved useful as a method for assessing genetic diversity in species of the Prunus genus. Key words: Prunus domestica, genetic diversity, Random Amplified Microsatellite markers (RAMs), deciduous.

Genetic diversity in Nordic and Baltic populations ofChondrostereum purpureum: a potential herbicide biocontrol agent

Summary We analysed genetic variation in the natural populations of a potential herbicide biocontrol agent, Chondrostereum purpureum, in Nordic and Baltic countries using random amplified microsatellite markers. The results showed high genetic diversity among the populations of this fungus, but almost a complete lack of local differentiation. The results implicate that any local strain from the area can be used as a biocontrol agent without a fear of introducing new genotypes to treatment areas.

Genetic variation in Polish strains of Gremmeniella abietina

SummaryThirty‐nine strains of Gremmeniella abietina were isolated between 1980 and 2005 from diseased or symptomless Pinus sylvestris and Pinus nigra, or sporadically Pinus jeffreyi and Pinus armandii from four regions of Poland with varying climatic conditions. Thirty‐five of the strains were genetically similar to the G. abietina A type found in Scandinavia, whereas four were similar to the B type. On the basis of random amplified microsatellite markers, very high intrapopulation and interpopulation genetic variation was detected among the strains. The average value of the Jaccard genetic similarity coefficients among strains was 0.56 for A type and 0.47 for B type, and the monomorphic loci partition was small, at 14% of all analysed loci for the A type. The degree of genetic variation of the fungal strains within each of the four regions was similar, showing no significant differences. The Jaccard coefficient for the strains isolated from P. sylvestris and P. nigra differed significantly, at 0.57 and 0.54 respectively. The genetic distance among A type strains from the investigated regions, expressed using Nei’s coefficient, was not correlated with geographic distance; however, it was highly dependent on elevation (correlation coefficient r = 0.92).

First Report of Phytophthora ramorum on Viburnum bodnantense in Belgium.

In the past decade, a new Phytophthora species inducing shoot canker on Rhododendron and dieback of Viburnum has been observed in Europe, mainly in Germany and the Netherlands, and California. This new pathogen has been named Phytophthora ramorum (3). In May 2002, a diseased Viburnum plant (Viburnum bodnantense) from the Plant Protection Service (Ministry of Agriculture, Belgium) was submitted to our laboratory for diagnosis. Symptoms included wilting, leaves turning from green to brown, discolored vascular tissues, and root necrosis. The plant came from a Belgian ornamental nursery that obtained supplies of stock plants from the Netherlands. Pieces of necrotic root tissue were excised, surface-disinfected, and transferred aseptically to a Phytophthora selective medium. P. ramorum was identified based on morphological characteristics, including the production of numerous, thin-walled chlamydospores (25 to 70 µm in diameter, average 43 µm) and deciduous, semi-papillate sporangia arranged in clusters. Radial growth after 6 days at 20°C on V8 juice agar was 2.8 mm per day. Random amplified microsatellite markers (RAMS) (2) from the total genomic DNA of the Belgian strain (CBS 110901) were similar to those of P. ramorum reference strains (CBS 101330, CBS 101332, and CBS 101554). Using PCR primers specific for P. ramorum, the identification was confirmed by W. A. Man in't Veld (Plantenziektenkundige Dienst, Wageningen, the Netherlands) (1). A pathogenicity test was carried out on three sterile cuttings of Rhododendron catawbiense (3). Brown lesions were observed on the inoculated cuttings after 6 to 7 days. None of the three uninoculated cuttings showed symptoms of infection. P. ramorum was reisolated from lesion margins on the inoculated cuttings. To our knowledge, this is the first report of the fungus from Belgium. Since our initial observation, we have found P. ramorum in other Belgian nurseries on R. yakusimanum.

Neofabraea populi in Hybrid Aspen Stands in Southern Finland

Symptoms typical of Neofabraea populi infection were identified from seven hybrid aspen (Populus tremula L. 2 P. tremuloides Michx.) stands in southern Finland. Morphology of the fungal isolates in vitro and ascospores released from apothecia on cankers was compared with reference isolates and the literature, respectively. A damage inventory was compiled in five stands. The percentages of healthy, diseased and dead stems per stand were calculated. The disease caused remarkable losses within stands (up to 53% mortality). Random amplified microsatellite markers were amplified from single-ascospore isolates, isolates from wood tissue and three reference isolates of N. populi. Both morphological and molecular analyses confirmed that the causal agent was N. populi, as suspected. The results imply that this fungus may become a major problem in hybrid aspen plantations in southern Finland.

Genetic Evidence for Somatic Haploidization in Developing Fruit Bodies of Armillaria tabescens

Armillaria spp. have vegetative hyphae with diploid uninucleate cells, but the fruit bodies of many species contain clamped dikaryotic hyphae. Earlier observations suggest that somatic haploidization takes place in developing fruit bodies. To verify this, a uninucleate diploid cell was isolated from each of the 49 mating combinations between single-spore isolates of Armillaria tabescens and they were fruited. Twenty-four isolates produced fruit bodies with at least a partially dikaryotic subhymenium. Dikaryotic hyphae were isolated from fruit-body primordia and homokaryons were obtained by micromanipulation or by protoplasting. Approximately half of the isolates proved to represent recombinant mating types in respect to parent homokaryons, and most of them contained recombinant haploid DNA, based on random-amplified microsatellite markers. The results show that the nuclei in dikaryotic hyphae found in fruit bodies result from somatic haploidization. The mechanism of haploidization remains unclear.

Genetic differentiation between European and North American populations of Phlebiopsis gigantea

We investigated the degree of genetic differentiation between geographically isolated populations of the saprophytic fungus Phlebiopsis gigantea from Europe and North America using two different molecular approaches: (i) denaturing gradient gel electrophoresis (DGGE) and sequence analyses of the ITS region, and (ii) genetic fingerprinting using random amplified microsatellite (RAMS) markers. The analysis included 84 homokaryotic single spore isolates of 28 European strains, 13 heterokaryotic strains from North America, and the Rotstop strain used as a biocontrol agent against Heterobasidion annosum. Three different ITS-alleles (designated as ITS/A, ITS/B and ITS/C) were separable by the DGGE, two of them occurring both in Europe and North America. However, sequence determination of these allele types showed that while the ITS/A alleles were identical between Europe and North America, the ITS/C alleles contained differences. Analysis of molecular variation (AMOVA) and neighbor joining (NJ) clustering analysis using altogether 28 RAMS markers revealed a considerable degree of differentiation between the European and North American populations. The results show that recent human-mediated P. gigantea migration between the continents is unlikely and that strains should not be transferred between continents for biocontrol purposes in order to avoid the introduction of novel genotypes.

Genetic Differentiation between European and North American Populations of Phlebiopsis gigantea

We investigated the degree of genetic differentiation between geographically isolated populations of the saprophytic fungus Phlebiopsis gigantea from Europe and North America using two different molecular approaches: (i) denaturing gradient gel electrophoresis (DGGE) and sequence analyses of the ITS region, and (ii) genetic fingerprinting using random amplified microsatellite (RAMS) markers. The analysis included 84 homokaryotic single spore isolates of 28 European strains, 13 heterokaryotic strains from North America, and the Rotstop strain used as a biocontrol agent against Heterobasidion annosum. Three different ITS-alleles (designated as ITS/A, ITS/B and ITS/C) were separable by the DGGE, two of them occurring both in Europe and North America. However, sequence determination of these allele types showed that while the ITS/A alleles were identical between Europe and North America, the ITS/C alleles contained differences. Analysis of molecular variation (AMOVA) and neighbor joining (NJ) clustering analysis using altogether 28 RAMS markers revealed a considerable degree of differentiation between the European and North American populations. The results show that recent human-mediated P. gigantea migration between the continents is unlikely and that strains should not be transferred between continents for biocontrol purposes in order to avoid the introduction of novel genotypes.

Genetic variation in Phlebiopsis gigantea as detected with random amplified microsatellite (RAMS) markers

Random Amplified Microsatellite (RAMS) markers were used to detect variation among Phlebiopsis gigantea isolates from seven European countries. The scoring of 26 bands revealed five markers common to all isolates and 21 variable markers, of which six occurred in only one strain. A considerable amount of variation was observed among the progeny isolates of single strains. The analysis of variable markers resulted in 75 different banding patterns among the 86 isolates studied. The high degree of polymorphism and heterozygosity observed in RAMS markers suggests that the amount of genetic variation within this fungus is considerable. The equal distribution of markers in strains from different locations and the lack of country-specific markers suggests that genetic differentiation among the populations is low. The lack of distinct groups of similar banding patterns supports the idea of P. gigantea being a true biological species consisting of a single intersterility group.