Descripción: Background: Prosopis alba (Fabaceae) is an important native tree adapted to arid and semiarid regions of north-western Argentina which is of great value as multipurpose species. Despite its importance, the genomic resources currently available for the entire Prosopis genus are still limited. Here we describe the development of a leaf transcriptome and the identification of new molecular markers that could support functional genetic studies in natural and domesticated populations of this genus.Results: Next generation DNA pyrosequencing technology applied to P. alba transcripts produced a total of 1,103,231 raw reads with an average length of 421 bp. De novo assembling generated a set of 15,814 isotigs and 71,101 non-assembled sequences (singletons) with an average of 991 bp and 288 bp respectively. A total of 39,000 unique singletons were identified after clustering natural and artificial duplicates from pyrosequencing reads.Regarding the non-redundant sequences or unigenes, 22,095 out of 54,814 were successfully annotated with Gene Ontology terms. Moreover, simple sequence repeats (SSRs) and single nucleotide polymorphisms (SNPs) were searched, resulting in 5,992 and 6,236 markers, respectively, throughout the genome. For the validation of the the predicted SSR markers, a subset of 87 SSRs selected through functional annotation evidence was successfully amplified from six DNA samples of seedlings. From this analysis, 11 of these 87 SSRs were identified as polymorphic. Additionally, another set of 123 nuclear polymorphic SSRs were determined in silico, of which 50% have the probability of being effectively polymorphic.Conclusions: This study generated a successful global analysis of the P. alba leaf transcriptome after bioinformatic and wet laboratory validations of RNA-Seq data.The limited set of molecular markers currently available will be significantly increased with the thousands of new markers that were identified in this study. This information will strongly contribute to genomics resources for P. alba functional analysis and genetics. Finally, it will also potentially contribute to the development of population-based genome studies in the genera. © 2013 Torales et al.; licensee BioMed Central Ltd.

Descripción: By using improved pulsed field gel electrophoresis conditions, the molecular karyotype of the reference clone CL Brener selected for Trypanosoma cruzi genome project was established. A total of 20 uniform chromosomal bands ranging in size from 0.45 to 3.5 Megabase pairs (Mbp) were resolved in a single run. The weighted sum of the chromosomal bands was approximately 87 Mbp. Chromoblots were hybridized with 39 different homologous probes, 13 of which identified single chromosomes. Several markers showed linkage and four different linkage groups were identified, each comprising two markers. Densitometric analysis suggests that most of the chromosomal bands contain two or more chromosomes representing either homologous chromosomes and/or heterologous chromosomes with similar sizes.

Descripción: Prosopis represents a valuable forest resource in arid and semiarid regions. Management of promising species requires information about genetic parameters, mainly the heritability (h2) of quantitative profitable traits. This parameter is traditionally estimated from progeny tests or half-sib analysis conducted in experimental stands. Such an approach estimates h 2 from the ratio of between-family/total phenotypic variance. These analyses are difficult to apply to natural populations of species with a long life cycle, overlapping generations, and a mixed mating system, without genealogical information. A promising alternative is the use of molecular marker information to infer relatedness between individuals and to estimate h 2 from the regression of phenotypic similarity on inferred relatedness. In the current study we compared h2 of 13 quantitative traits estimated by these two methods in an experimental stand of P. alba, where genealogical information was available. We inferred pairwise relatedness by Ritland's method using six microsatellite loci. Relatedness and heritability estimates from molecular information were highly correlated to the values obtained from genealogical data. Although Ritland's method yields lower h 2 estimates and tends to overestimate genetic correlations between traits, this approach is useful to predict the expected relative gain of different quantitative traits under selection without genealogical information.

Descripción: The subcellular localization and physiological functions of biomolecules are closely related and thus it is crucial to precisely determine the distribution of different molecules inside the intracellular structures. This is frequently accomplished by fluorescence microscopy with well-characterized markers and posterior evaluation of the signal colocalization. Rigorous study of colocalization requires statistical analysis of the data, albeit yet no single technique has been established as a standard method. Indeed, the few methods currently available are only accurate in images with particular characteristics. Here, we introduce a new algorithm to automatically obtain the true colocalization between images that is suitable for a wide variety of biological situations. To proceed, the algorithm contemplates the individual contribution of each pixel's fluorescence intensity in a pair of images to the overall Pearsońs correlation and Manders' overlap coefficients. The accuracy and reliability of the algorithm was validated on both simulated and real images that reflected the characteristics of a range of biological samples. We used this algorithm in combination with image restoration by deconvolution and time-lapse confocal microscopy to address the localization of MEK1 in the mitochondria of different cell lines. Appraising the previously described behavior of Akt1 corroborated the reliability of the combined use of these techniques. Together, the present work provides a novel statistical approach to accurately and reliably determine the colocalization in a variety of biological images. © 2011 Villalta et al.

Descripción: Background: Freshwater lymnaeid snails can act as the intermediate hosts for trematode parasites such as the liver fluke Fasciola hepatica, that cause significant economic and biomedical burden worldwide, particularly through bovine fascioliasis. Transmission potential is tightly coupled to local compatibility with snail hosts, so accurate identification of lymnaeid species is crucial for understanding disease risk, especially when invasive species are encountered. Mendoza Province, in Argentina, is a center of livestock production and also an area of endemic fascioliasis transmission. However, the distribution of lymnaeid species in the region is not well known. Methods. This study examined lymnaeid snails from seven localities in the Department of Malarguë, Mendoza Province, using morphological and molecular analyses and also describing ecological variables associated with snail presence. Results: While morphological characters identified two species of lymnaeid, Galba truncatula and G. viatrix, molecular data revealed a third, cryptic species, G. neotropica, which was sympatric with G. viatrix. G. truncatula was exclusively found in high altitude (>1900 meters above sea level [masl]) sites, whereas mixed G. neotropica/G. viatrix localities were at middle elevations (1300-1900 masl), and G. viatrix was found alone at the lowest altitude sites (<1300 masl). Phylogenetic analysis using two mitochondrial markers revealed G. neotropica and G. viatrix to be closely related, and given their morphological similarities, their validities as separate taxonomic entities should be questioned. Conclusions: This study highlights the need of a robust taxonomic framework for the identification of lymnaeid snails, incorporating molecular, morphological and ecological variables while avoiding nomenclature redundancy. As the three species observed here, including one alien invasive species, are considered hosts of varying susceptibility to Fasciola parasites, and given the economic importance of fascioliasis for livestock production, this research has critical importance for the ultimate aim of controlling disease transmission. © 2013 Standley et al.; licensee BioMed Central Ltd.

Descripción: Sequence analysis of the internal transcribed spacer (ITS) of the 18S(ITS1)-5.8S-26S(ITS2) rDNA region was performed in order to analyse the phylogenetic relationships between 13 Patagonian species of the genus Berberis (Berberidaceae). The divergence values between the pairwise sequence in the studied Patagonian species were in the range 2.9-22.9%. The lengths of the ITS1 and ITS2 sequences were in the range 227-231 bp and 220-224 bp, respectively, and the 5.8S sequence was 159 bp throughout all species. B. microphylla sensu Landrum does not appear to be monophyletic based on current sampling. Indeed, we suggest that B. microphylla should be distinguished from B. buxifolia, B. parodii, and B. heterophylla. ITS sequences, together with data obtained from morphological, biochemical, amplified fragment length polymorphism, and cytological characterizations, support the existence of diploid and polyploid hybrid speciation in the genus. © 2007 The Linnean Society of London.

Descripción: The proopiomelanocortin gene (POMC) is expressed in a group of neurons present in the arcuate nucleus of the hypothalamus. Neuron-specific POMC expression in mammals is conveyed by two distal enhancers, named nPE1 and nPE2. Previous transgenic mouse studies showed that nPE1 and nPE2 independently drive reporter gene expression to POMC neurons. Here, we investigated the evolutionary mechanisms that shaped not one but two neuron- specific POMC enhancers and tested whether nPE1 and nPE2 drive identical or complementary spatiotemporal expression patterns. Sequence comparison among representative genomes of most vertebrate classes and mammalian orders showed that nPE1 is a placental novelty. Using in silico paleogenomics we found that nPE1 originated from the exaptation of a mammalian- apparent LTR retrotransposon sometime between the metatherian/ eutherian split (147 Mya) and the placental mammal radiation (≈90 Mya). Thus, the evolutionary origin of nPE1 differs, in kind and time, from that previously demonstrated for nPE2, which was exapted from a CORE-short interspersed nucleotide element (SINE) retroposon before the origin of prototherians, 166 Mya. Transgenic mice expressing the fluorescent markers tomato and EGFP driven by nPE1 or nPE2, respectively, demonstrated coexpression of both reporter genes along the entire arcuate nucleus. The onset of reporter gene expression guided by nPE1 and nPE2 was also identical and coincidental with the onset of Pomc expression in the presumptive mouse diencephalon. Thus, the independent exaptation of two unrelated retroposons into functional analogs regulating neuronal POMC expression constitutes an authentic example of convergent molecular evolution of cell-specific enhancers.

Descripción: DNA size polymorphisms were utilized in a study of 24 natural populations of Ceratitis capitata Wiedemann (Diptera: Tephritidae) from Argentina. The first intron of alcohol dehydrogenase 1 gene (Adh1) was amplified using exon priming intron crossing-polymerase chain reaction. Three size variants were detected among the 307 samples analyzed. To better differentiate the size variants, further digestion of PCR products with the EcoRI restriction enzyme was carried out. Complete nucleotide sequences of the three-allele variants were obtained and single changes, insertions, deletions, and EcoRI recognition sites were located. Population allele frequencies were analyzed and a global mean heterozygosity (He) of 0.33 was obtained. In most populations, observed allelic frequencies conformed to Hardy-Weinberg expectations. Significant differences between provinces and sampling sites within these provinces, and among some populations were found. The average number of insects exchanged among populations (Nm) was estimated and high values were observed between Argentina and populations from two African countries (Morocco and Kenya), Australia, and Hawaii (Kauai). Pest introduction sources and dispersion patterns in Argentina are discussed based on these results as well as on available bibliographical data. © 2010 The Authors. Journal compilation © 2010 The Netherlands Entomological Society.

Descripción: Tyrant flycatchers (Aves: Tyrannidae) are endemic to the New World, and many species of this group are threatened or near-threatened at the global level. The aim of this study was to test the 18 microsatellite markers that have been published for other Tyrant flycatchers in the Strange-tailed Tyrant (Alectrurus risora) and the Sharp-tailed Tyrant (Culicivora caudacuta), two endemic species of southern South American grasslands that are classified as vulnerable. We also analyzed the usefulness of loci in relation to phylogenetic distance to the source species. Amplification success was high in both species (77 to 83%) and did not differ between the more closely and more distantly related species to the source species. Polymorphism success was also similar for both species, with 9 and 8 loci being polymorphic, respectively. An increased phylogenetic distance thus does not gradually lead to allelic or locus dropouts, implying that in Tyrant flycatchers, the published loci are useful independent of species relatedness. © FUNPEC-RP.

Descripción: Munida gregaria and M. subrugosa have been considered two different species for more than a century; however, after a recent molecular phylogenetic study, they are considered a single polymorphic species. Yet, the use of markers to diagnose species may be misleading when divergence between species is recent, since a speciation event may be obscured by the retention and stochastic sorting of ancestral polymorphisms. The morphs gregaria and subrugosa of Munida gregaria constitute an interesting case for the study of behavioural isolation since they are sympatric, breed at the same time of the year, and might have experienced a recent speciation. Mating behaviour observations and mate choice mating trials were conducted in order to investigate the potential existence of a behavioural prezygotic barrier to gene flow between these two morphs. Since factors involved in mate choice in galatheids are unknown, the four possible combinations of the two different morphs in trios were used to test for the existence of mate choice. Video recordings of all the possible trio combinations revealed that there was cross-attraction between males and females of different morphs. Females bearing partial broods participated in encounters as well as non-ovigerous females. The frequency and duration of homo- and heterotypic encounters were registered, and a reproductive isolation index was calculated for each variable for each trio. The isolation indexes calculated were not different from zero indicating random mating, and were not affected by the composition of the trio or the partial ovigerous condition of females. These results provided evidence of the absence of behavioural prezygotic barriers to gene flow between the morphs gregaria and subrugosa of M. gregaria. © 2011 Springer-Verlag and AWI.

Descripción: Background: Subtractive hybridization methods are valuable tools for identifying differentially regulated genes in a given tissue avoiding redundant sequencing of clones representing the same expressed genes, maximizing detection of low abundant transcripts and thus, affecting the efficiency and cost effectiveness of small scale cDNA sequencing projects aimed to the specific identification of useful genes for breeding purposes. The objective of this work is to evaluate alternative strategies to high-throughput sequencing projects for the identification of novel genes differentially expressed in sunflower as a source of organ-specific genetic markers that can be functionally associated to important traits. Results: Differential organ-specific ESTs were generated from leaf, stem, root and flower bud at two developmental stages (R1 and R4). The use of different sources of RNA as tester and driver cDNA for the construction of differential libraries was evaluated as a tool for detection of rare or low abundant transcripts. Organ-specificity ranged from 75 to 100% of non-redundant sequences in the different cDNA libraries. Sequence redundancy varied according to the target and driver cDNA used in each case. The R4 flower cDNA library was the less redundant library with 62% of unique sequences. Out of a total of 919 sequences that were edited and annotated, 318 were non-redundant sequences. Comparison against sequences in public databases showed that 60% of non-redundant sequences showed significant similarity to known sequences. The number of predicted novel genes varied among the different cDNA libraries, ranging from 56% in the R4 flower to 16 % in the R1 flower bud library. Comparison with sunflower ESTs on public databases showed that 197 of non-redundant sequences (60%) did not exhibit significant similarity to previously reported sunflower ESTs. This approach helped to successfully isolate a significant number of new reported sequences putatively related to responses to important agronomic traits and key regulatory and physiological genes. Conclusions: The application of suppressed subtracted hybridization technology not only enabled the cost effective isolation of differentially expressed sequences but it also allowed the identification of novel sequences in sunflower from a relative small number of analyzed sequences when compared to major sequencing projects. © 2003 Fernández et al; licensee BioMed Central Ltd.