SNP identification, verification, and utility for population genetics in a non-model genus

<p>Abstract</p> <p>Background</p> <p>By targeting SNPs contained in both coding and non-coding areas of the genome, we are able to identify genetic differences and characterize genome-wide patterns of variation among individuals, populations and species. We investigated the utility of 454 sequencing and MassARRAY genotyping for population genetics in natural populations of the teleost, <it>Fundulus heteroclitus </it>as well as closely related <it>Fundulus </it>species (<it>F. grandis</it>, <it>F. majalis </it>and <it>F. similis</it>).</p> <p>Results</p> <p>We used 454 pyrosequencing and MassARRAY genotyping technology to identify and type 458 genome-wide SNPs and determine genetic differentiation within and between populations and species of <it>Fundulus</it>. Specifically, pyrosequencing identified 96 putative SNPs across coding and non-coding regions of the <it>F. heteroclitus </it>genome: 88.8% were verified as true SNPs with MassARRAY. Additionally, putative SNPs identified in <it>F. heteroclitus </it>EST sequences were verified in most (86.5%) <it>F. heteroclitus </it>individuals; fewer were genotyped in <it>F. grandis </it>(74.4%), <it>F. majalis </it>(72.9%), and <it>F. similis </it>(60.7%) individuals. SNPs were polymorphic and showed latitudinal clinal variation separating northern and southern populations and established isolation by distance in <it>F. heteroclitus </it>populations. In <it>F. grandis</it>, SNPs were less polymorphic but still established isolation by distance. Markers differentiated species and populations.</p> <p>Conclusions</p> <p>In total, these approaches were used to quickly determine differences within the <it>Fundulus </it>genome and provide markers for population genetic studies.</p

Incompletitud corporal en la persona posamputada portadora de dolor de miembro fantasma: estudio cualitativo

Introducción: En el cuerpo se vivencía el mundo, la alegría, el encuentro, la seguridad, el dolor las reacciones anatomofisiológicas inherentemente al cuerpo; se experimenta la afección por el dolor físico y psíquico, con sensaciones corporales de ansiedad, temor y angustia. Materiales y Métodos: Estudio de abordaje cualitativo-descriptivo, exploratorio; como técnica se empleó la entrevista semiestructurada, de los informantes seleccionados se trataba de personas que fueron atendidas en el Hospital General de Durango, México; participaron 12 personas que al ser amputadas debutaron con dolor de miembro fantasma; el soporte teórico estuvo apoyado por la corporeidad de Le Breton. Resultados y Discusión: En la persona con amputación, la relación de él con el mundo se altera, ya que la amputación cambia no sólo la forma de verse a sí mismo, sino también la forma de interactuar con su entorno, su familia, amigos, compañeros y sociedad en general. Conclusiones: Las enfermeras deben fortalecer la relación enfermera/persona para vivir un encuentro real mostrando actitud de empatía, estar siempre ayudando y apoyando. Igualmente, escuchando, comprendiendo y educando, las personas al sentirse cuidadas contribuirán para que sean tratadas como un ser completo y harán que en su cotidiano la experiencia de vivir con el dolor de miembro fantasma se transforme en una condición aceptable y le permita enfrentar mejor su situación.Cómo citar este artículo: Ochoa MC, Bustamante S, Hernández C. Incompletitud corporal en la persona posamputada portadora de dolor de miembro fantasma: estudio cualitativo. Rev Cuid. 2015; 6(1): 941-6. http://dx.doi.org/10.15649/cuidarte.v6i1.14

Evaluation and Characterization of Post-Stroke Lung Damage in a Murine Model of Cerebral Ischemia

After stroke and other brain injuries, there is a high incidence of respiratory complications such as pneumonia or acute lung injury. The molecular mechanisms that drive the brain-lung interaction post-stroke have not yet been elucidated. We performed transient middle cerebral artery occlusion (MCAO) and sham surgery on C57BL/6J mice and collected bronchoalveolar lavage fluid (BALF), serum, brain, and lung homogenate samples 24 h after surgery. A 92 proteins-panel developed by Olink Proteomics ® was used to analyze the content in BALF and lung homogenates. MCAO animals had higher protein concentration levels in BALF than sham -controls, but these levels did not correlate with the infarct volume. No alteration in alveolar-capillary barrier permeability was observed. A total of 12 and 14 proteins were differentially expressed between the groups (FDR < 0.1) in BALF and lung tissue homogenates, respectively. Of those, HGF, TGF-α, and CCL2 were identified as the most relevant to this study. Their protein expression patterns were verified by ELISA. This study confirmed that post-stroke lung damage was not associated with increased lung permeability or cerebral ischemia severity. Furthermore, the dysregulation of HGF, TGF-α, and CCL2 in BALF and lung tissue after ischemia could play an important role in the molecular mechanisms underlying stroke-induced lung damage

Kinesin Is an Evolutionarily Fine-Tuned Molecular Ratchet-and-Pawl Device of Decisively Locked Direction

Conventional kinesin is a dimeric motor protein that transports membranous organelles toward the plus-end of microtubules (MTs). Individual kinesin dimers show steadfast directionality and hundreds of consecutive steps, yetthe detailed physical mechanism remains unclear. Here we compute free energies for the entire dimer-MT system for all possible interacting configurations by taking full account of molecular details. Employing merely first principles and several measured binding and barrier energies, the system-level analysis reveals insurmountable energy gaps between configurations, asymmetric ground state caused by mechanically lifted configurational degeneracy, and forbidden transitions ensuring coordination between both motor domains for alternating catalysis. This wealth of physical effects converts a kinesin dimer into a molecular ratchet-and-pawl device, which determinedly locks the dimer's movement into the MT plus-end and ensures consecutive steps in hand-over-hand gait.Under a certain range of extreme loads, however, the ratchet-and-pawl device becomes defective but not entirely abolished to allow consecutive back-steps. This study yielded quantitative evidence that kinesin's multiple molecular properties have been evolutionarily adapted to fine-tune the ratchet-and-pawl device so as to ensure the motor's distinguished performance.Comment: 10 printed page

Population genomic analysis reveals a rich speciation and demographic history of orang-utans (Pongo pygmaeus and Pongo abelii)

To gain insights into evolutionary forces that have shaped the history of Bornean and Sumatran populations of orang-utans, we compare patterns of variation across more than 11 million single nucleotide polymorphisms found by previous mitochondrial and autosomal genome sequencing of 10 wild-caught orang-utans. Our analysis of the mitochondrial data yields a far more ancient split time between the two populations (~3.4 million years ago) than estimates based on autosomal data (0.4 million years ago), suggesting a complex speciation process with moderate levels of primarily male migration. We find that the distribution of selection coefficients consistent with the observed frequency spectrum of autosomal non-synonymous polymorphisms in orang-utans is similar to the distribution in humans. Our analysis indicates that 35% of genes have evolved under detectable negative selection. Overall, our findings suggest that purifying natural selection, genetic drift, and a complex demographic history are the dominant drivers of genome evolution for the two orang-utan populations.Publisher PDFPeer reviewe

Mcidas mutant mice reveal a two-step process for the specification and differentiation of multiciliated cells in mammals

Motile cilia on multiciliated cells (MCCs) function in fluid clearance over epithelia. Studies with Xenopus embryos and individuals with the congenital respiratory disorder reduced generation of multiple motile cilia (RGMC), have implicated the nuclear protein MCIDAS (MCI), in the transcriptional regulation of MCC specification and differentiation. Recently, a paralogous protein, geminin coiled-coil domain containing (GMNC), was also shown to be required for MCC formation. Surprisingly, in contrast to the presently held view, we find that Mci mutant mice can specify MCC precursors. However, these precursors cannot produce multiple basal bodies, and mature into single ciliated cells. We identify an essential role for MCI in inducing deuterosome pathway components for the production of multiple basal bodies. Moreover, GMNC and MCI associate differentially with the cell-cycle regulators E2F4 and E2F5, which enables them to activate distinct sets of target genes (ciliary transcription factor genes versus basal body amplification genes). Our data establish a previously unrecognized two-step model for MCC development: GMNC functions in the initial step for MCC precursor specification. GMNC induces Mci expression that drives the second step of basal body production for multiciliation

Knockout of the dhfr-ts Gene in Trypanosoma cruzi Generates Attenuated Parasites Able to Confer Protection against a Virulent Challenge

Chagas disease is the clinical manifestation of the infection produced by the flagellate parasite Trypanosoma cruzi and currently there is no vaccine to prevent this disease. Therefore, different approaches or alternatives are urgently needed. Vaccination with live attenuated parasites has been used effectively in mice to reduce parasitemia and histological damage. However, the use of live parasites as inmunogens is controversial due to the risk of reversion to a virulent phenotype. In this work we genetically manipulated a naturally attenuated strain of T. cruzi in order to produce parasites with impaired replication and infectivity, using the mutation as a safety device against reversion to virulence. We show that genetically modified parasites display a lower proliferation rate in vitro and induced almost undetectable levels of T. cruzi specific CD8+ T cells when injected in mice. Furthermore, the immune response induced by these live mutant parasites confers protection against a subsequent virulent infection even a year after the original immunization

Drug discovery for Chagas disease should consider Trypanosoma cruzi strain diversity.

This opinion piece presents an approach to standardisation of an important aspect of Chagas disease drug discovery and development: selecting Trypanosoma cruzi strains for in vitro screening. We discuss the rationale for strain selection representing T. cruzi diversity and provide recommendations on the preferred parasite stage for drug discovery, T. cruzi discrete typing units to include in the panel of strains and the number of strains/clones for primary screens and lead compounds. We also consider experimental approaches for in vitro drug assays. The Figure illustrates the current Chagas disease drug-discovery and development landscape

Effect of Roscovitine on nuclear maturation, MPF and MAP kinase activity and embryo development of prepubertal goat oocytes

The low number of embryos obtained from IVM-IVF-IVC of prepubertal goat oocytes could be due to an incomplete cytoplasmic maturation. Roscovitine (ROS) inhibits MPF and MAP kinase activity and maintains the oocyte at Germinal Vesicle (GV) stage. The aim of this study was to determine if meiotic activity is arrested in prepubertal goat oocytes cultured with 0, 12.5, 25, 50 and 100 m M of ROS for 24 h. A group of oocytes from adult goats was cultured with 25 m M of ROS to compare the effect of ROS on prepubertal and adult goat oocytes. A sample of oocytes was stained to evaluate the nuclear stage at oocyte collection time and after ROS incubation. IVM-oocytes not exposed to ROS formed the control group. Prepubertal goat IVM-oocytes were inseminated and cultured for 8 days. The percentage of oocytes at GV stage, after exposition to ROS was significantly higher in adult goat oocytes (64.5%) than in prepubertal goat oocytes. No differences were found among 25, 50 and 100 m M ROS concentrations (29, 23 and 26%, oocytes at GV stage, respectively). After 8 days of culture, no differences in total embryos were observed between control oocytes and oocytes treated with 12.5 and 25 m M (45.2, 36.1 and 39.4%, respectively), however the percentage of lastocysts was higher in the control group. Western blot for the MAPK and p34 cdc2 showed that both enzymes were active in prepubertal goat oocytes after 24 h of ROS exposition. In conclusion, a low percentage of prepubertal goat oocytes reached GV stage after ROS incubation; possibly because most of them had reinitiated the meiosis inside the follicle. ROS did not affect fertilization or total embryos but ROS showed a negative effect on blastocyst development