HYTEST Phase I Facility Commissioning and Modeling

The purpose of this document is to report the first year accomplishments of two coordinated Laboratory Directed Research and Development (LDRD) projects that utilize a hybrid energy testing laboratory that couples various reactors to investigate system reactance behavior. This work is the first phase of a series of hybrid energy research and testing stations - referred to hereafter as HYTEST facilities – that are planned for construction and operation at the Idaho National Laboratory (INL). A HYTEST Phase I facility was set up and commissioned in Bay 9 of the Bonneville County Technology Center (BCTC). The purpose of this facility is to utilize the hydrogen and oxygen that is produced by the High Temperature Steam Electrolysis test reactors operating in Bay 9 to support the investigation of kinetic phenomena and transient response of integrated reactor components. This facility provides a convenient scale for conducting scoping tests of new reaction concepts, materials performance, new instruments, and real-time data collection and manipulation for advance process controls. An enclosed reactor module was assembled and connected to a new ventilation system equipped with a variable-speed exhaust blower to mitigate hazardous gas exposures, as well as contract with hot surfaces. The module was equipped with a hydrogen gas pump and receiver tank to supply high quality hydrogen to chemical reactors located in the hood

Phylogenetically Widespread Polyembryony in Cyclostome Bryozoans and the Protracted Asynchronous Release of Clonal Brood-Mates

The file attached is the Published/publisher’s pdf version of the articl

Physical activity, sedentary time and physical capability in early old age: British birth cohort study.

PURPOSE: To investigate the associations of time spent sedentary, in moderate-to-vigorous-intensity physical activity (MVPA) and physical activity energy expenditure (PAEE) with physical capability measures at age 60-64 years. METHODS: Time spent sedentary and in MVPA and, PAEE were assessed using individually calibrated combined heart rate and movement sensing among 1727 participants from the MRC National Survey of Health and Development in England, Scotland and Wales as part of a detailed clinical assessment undertaken in 2006-2010. Multivariable linear regression models were used to examine the cross-sectional associations between standardised measures of each of these behavioural variables with grip strength, chair rise and timed up-&-go (TUG) speed and standing balance time. RESULTS: Greater time spent in MVPA was associated with higher levels of physical capability; adjusted mean differences in each capability measure per 1 standard deviation increase in MVPA time were: grip strength (0.477 kg, 95% confidence interval (CI): 0.015 to 0.939), chair rise speed (0.429 stands/min, 95% CI: 0.093 to 0.764), standing balance time (0.028 s, 95% CI: 0.003 to 0.053) and TUG speed (0.019 m/s, 95% CI: 0.011 to 0.026). In contrast, time spent sedentary was associated with lower grip strength (-0.540 kg, 95% CI: -1.013 to -0.066) and TUG speed (-0.011 m/s, 95% CI: -0.019 to -0.004). Associations for PAEE were similar to those for MVPA. CONCLUSION: Higher levels of MVPA and overall physical activity (PAEE) are associated with greater levels of physical capability whereas time spent sedentary is associated with lower levels of capability. Future intervention studies in older adults should focus on both the promotion of physical activity and reduction in time spent sedentary.This work was supported by the UK Medical Research Council (U120063239, U123092720, MC_UU_12019/1, MC_UU_12019/4, MC_UU_12015/3, and MC_UU_12015/4).This is the final published version. It first appeared at http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0126465

Addressing statistical biases in nucleotide-derived protein databases for proteogenomic search strategies

[Image: see text] Proteogenomics has the potential to advance genome annotation through high quality peptide identifications derived from mass spectrometry experiments, which demonstrate a given gene or isoform is expressed and translated at the protein level. This can advance our understanding of genome function, discovering novel genes and gene structure that have not yet been identified or validated. Because of the high-throughput shotgun nature of most proteomics experiments, it is essential to carefully control for false positives and prevent any potential misannotation. A number of statistical procedures to deal with this are in wide use in proteomics, calculating false discovery rate (FDR) and posterior error probability (PEP) values for groups and individual peptide spectrum matches (PSMs). These methods control for multiple testing and exploit decoy databases to estimate statistical significance. Here, we show that database choice has a major effect on these confidence estimates leading to significant differences in the number of PSMs reported. We note that standard target:decoy approaches using six-frame translations of nucleotide sequences, such as assembled transcriptome data, apparently underestimate the confidence assigned to the PSMs. The source of this error stems from the inflated and unusual nature of the six-frame database, where for every target sequence there exists five “incorrect” targets that are unlikely to code for protein. The attendant FDR and PEP estimates lead to fewer accepted PSMs at fixed thresholds, and we show that this effect is a product of the database and statistical modeling and not the search engine. A variety of approaches to limit database size and remove noncoding target sequences are examined and discussed in terms of the altered statistical estimates generated and PSMs reported. These results are of importance to groups carrying out proteogenomics, aiming to maximize the validation and discovery of gene structure in sequenced genomes, while still controlling for false positives

Assessing the digenic model in rare disorders using population sequencing data

An important fraction of patients with rare disorders remains with no clear genetic diagnostic, even after whole-exome or whole-genome sequencing, posing a difficulty in giving adequate treatment and genetic counseling. The analysis of genomic data in rare disorders mostly considers the presence of single gene variants in coding regions that follow a concrete monogenic mode of inheritance. A digenic inheritance, with variants in two functionally-related genes in the same individual, is a plausible alternative that might explain the genetic basis of the disease in some cases. In this case, digenic disease combinations should be absent or underrepresented in healthy individuals. We develop a framework to evaluate the significance of digenic combinations and test its statistical power in different scenarios. We suggest that this approach will be relevant with the advent of new sequencing efforts including hundreds of thousands of samples

Identification of 4 novel human ocular coloboma genes ANK3, BMPR1B, PDGFRA, and CDH4 through evolutionary conserved vertebrate gene analysis

Purpose: Ocular coloboma arises from genetic or environmental perturbations that inhibit optic fissure (OF) fusion during early eye development. Despite high genetic heterogeneity, 70% to 85% of patients remain molecularly undiagnosed. In this study, we have identified new potential causative genes using cross-species comparative meta-analysis. Methods: Evolutionarily conserved differentially expressed genes were identified through in silico analysis, with in situ hybridization, gene knockdown, and rescue performed to confirm spatiotemporal gene expression and phenotype. Interrogation of the 100,000 Genomes Project for putative pathogenic variants was performed. Results: Nine conserved differentially expressed genes between zebrafish and mouse were identified. Expression of zebrafish ank3a, bmpr1ba/b, cdh4, and pdgfaa was localized to the OF, periocular mesenchyme cells, or ciliary marginal zone, regions traversed by the OF. Knockdown of ank3, bmpr1b, and pdgfaa revealed a coloboma and/or microphthalmia phenotype. Novel pathogenic variants in ANK3, BMPR1B, PDGFRA, and CDH4 were identified in 8 unrelated coloboma families. We showed BMPR1B rescued the knockdown phenotype but variant messenger RNAs failed, providing evidence of pathogenicity. Conclusion: We show the utility of cross-species meta-analysis to identify several novel coloboma disease-causing genes. There is a potential to increase the diagnostic yield for new and unsolved patients while adding to our understanding of the genetic basis of OF morphogenesis

Author Correction: Nuclear-mitochondrial DNA segments resemble paternally inherited mitochondrial DNA in humans

An amendment to this paper has been published and can be accessed via a link at the top of the paper

DYNC2H1 hypomorphic or retina-predominant variants cause nonsyndromic retinal degeneration

Purpose: Determining the role of DYNC2H1 variants in nonsyndromic inherited retinal disease (IRD). Methods: Genome and exome sequencing were performed for five unrelated cases of IRD with no identified variant. In vitro assays were developed to validate the variants identified (fibroblast assay, induced pluripotent stem cell [iPSC] derived retinal organoids, and a dynein motility assay). Results: Four novel DYNC2H1 variants (V1, g.103327020_103327021dup; V2, g.103055779A>T; V3, g.103112272C>G; V4, g.103070104A>C) and one previously reported variant (V5, g.103339363T>G) were identified. In proband 1 (V1/V2), V1 was predicted to introduce a premature termination codon (PTC), whereas V2 disrupted the exon 41 splice donor site causing incomplete skipping of exon 41. V1 and V2 impaired dynein-2 motility in vitro and perturbed IFT88 distribution within cilia. V3, homozygous in probands 2–4, is predicted to cause a PTC in a retina-predominant transcript. Analysis of retinal organoids showed that this new transcript expression increased with organoid differentiation. V4, a novel missense variant, was in trans with V5, previously associated with Jeune asphyxiating thoracic dystrophy (JATD). Conclusion: The DYNC2H1 variants discussed herein were either hypomorphic or affecting a retina-predominant transcript and caused nonsyndromic IRD. Dynein variants, specifically DYNC2H1 variants are reported as a cause of non syndromic IRD

Cross-oncopanel study reveals high sensitivity and accuracy with overall analytical performance depending on genomic regions

BackgroundTargeted sequencing using oncopanels requires comprehensive assessments of accuracy and detection sensitivity to ensure analytical validity. By employing reference materials characterized by the U.S. Food and Drug Administration-led SEquence Quality Control project phase2 (SEQC2) effort, we perform a cross-platform multi-lab evaluation of eight Pan-Cancer panels to assess best practices for oncopanel sequencing.ResultsAll panels demonstrate high sensitivity across targeted high-confidence coding regions and variant types for the variants previously verified to have variant allele frequency (VAF) in the 5-20% range. Sensitivity is reduced by utilizing VAF thresholds due to inherent variability in VAF measurements. Enforcing a VAF threshold for reporting has a positive impact on reducing false positive calls. Importantly, the false positive rate is found to be significantly higher outside the high-confidence coding regions, resulting in lower reproducibility. Thus, region restriction and VAF thresholds lead to low relative technical variability in estimating promising biomarkers and tumor mutational burden.ConclusionThis comprehensive study provides actionable guidelines for oncopanel sequencing and clear evidence that supports a simplified approach to assess the analytical performance of oncopanels. It will facilitate the rapid implementation, validation, and quality control of oncopanels in clinical use.Peer reviewe

Mechanical Influences on Morphogenesis of the Knee Joint Revealed through Morphological, Molecular and Computational Analysis of Immobilised Embryos

Very little is known about the regulation of morphogenesis in synovial joints. Mechanical forces generated from muscle contractions are required for normal development of several aspects of normal skeletogenesis. Here we show that biophysical stimuli generated by muscle contractions impact multiple events during chick knee joint morphogenesis influencing differential growth of the skeletal rudiment epiphyses and patterning of the emerging tissues in the joint interzone. Immobilisation of chick embryos was achieved through treatment with the neuromuscular blocking agent Decamethonium Bromide. The effects on development of the knee joint were examined using a combination of computational modelling to predict alterations in biophysical stimuli, detailed morphometric analysis of 3D digital representations, cell proliferation assays and in situ hybridisation to examine the expression of a selected panel of genes known to regulate joint development. This work revealed the precise changes to shape, particularly in the distal femur, that occur in an altered mechanical environment, corresponding to predicted changes in the spatial and dynamic patterns of mechanical stimuli and region specific changes in cell proliferation rates. In addition, we show altered patterning of the emerging tissues of the joint interzone with the loss of clearly defined and organised cell territories revealed by loss of characteristic interzone gene expression and abnormal expression of cartilage markers. This work shows that local dynamic patterns of biophysical stimuli generated from muscle contractions in the embryo act as a source of positional information guiding patterning and morphogenesis of the developing knee joint