bluuebunny/crossref_metadata_2025_split huggingface dataset:
Employing single-stranded DNA donors for the high-throughput production of conditional knockout alleles in mice
Authors: Lanza Denise G. , Gaspero Angelina , Lorenzo Isabel , Liao Lan , Zheng Ping , Wang Ying , Deng Yu , Cheng Chonghui , Zhang Chuansheng , Rasband Matthew N. , Seavitt John R. , DeMayo Francisco J. , Xu Jianming , Dickinson Mary E. , Beaudet Arthur L. , Heaney Jason D. 
Published: September 2017

ABSTRACTThe International Mouse Phenotyping Consortium is generating null allele mice for every protein-coding gene in the genome and characterizing these mice to identify gene-phenotype associations. To test the feasibility of using CRISPR/Cas9 gene editing to generate conditional knockout mice for this large-scale resource, we employed Cas9-mediated homology driven repair (HDR) with short and long single-stranded oligodeoxynucleotides (ssODNs and lssODNs). Using pairs of guides and ssODNs donating loxP sites, we obtained putative conditional allele founder mice, harboring both loxP sites, for 23 of 30 genes targeted. LoxP sites integratedin cisin at least one F0 for 18 of 23 targeted genes. However, loxP sites were mutagenized in 4 of 18in cislines. HDR efficiency correlated with Cas9 cutting efficiency but was not influenced by ssODN homology arm symmetry. By contrast, using pairs of guides and a single lssODN to introduce a loxP-flanked exon, conditional allele founders were generated for all 4 genes targeted. Our studies demonstrate that Cas9-mediated HDR with pairs of ssODNs can generate conditional null alleles at many loci, but reveal inefficiencies when applied at scale. In contrast, lssODNs are amenable to high-throughput production of conditional alleles when they can be employed.
High-throughput creation and functional profiling of eukaryotic DNA sequence variant libraries using CRISPR/Cas9
Authors: Guo Xiaoge , Chavez Alejandro , Tung Angela , Chan Yingleong , Cecchi Ryan , Garnier Santiago Lopez , Kaas Christian , Kelsic Eric , Schubert Max , DiCarlo James E. , Collins James J. , Church George M. 
Published: September 2017

ABSTRACTConstruction of genetic variant libraries with phenotypic measurement is central to advancing today’s functional genomics, and remains a grand challenge. Here, we introduce a Cas9-based approach for generating pools of mutants with defined genetic alterations (deletions, substitutions and insertions), along with methods for tracking their fitnessen masse. We demonstrate the utility of our approach in performing focused analysis of hundreds of mutants of a single protein and in investigating the biological function of an entire family of poorly characterized genetic elements. Our platform allows fundamental biology questions to be investigated in a quick, easy and affordable manner.
PHENOS: a high-throughput and flexible tool for microorganism growth phenotyping on solid media
Authors: Barton David B. H. , Georghiou Danae , Dave Neelam , Alghamdi Majed , Walsh Thomas A. , Louis Edward J. , Foster Steven S. 
Published: September 2017

ABSTRACTBACKGROUNDMicrobial arrays, with a large number of different strains on a single plate printed with robotic precision, underpin an increasing number of genetic and genomic approaches. These include Synthetic Genetic Array analysis, high-throughput Quantitative Trait Loci (QTL) analysis and 2-hybrid techniques. Measuring the growth of individual colonies within these arrays is an essential part of many of these techniques but is useful for any work with arrays. Measurement is typically done using intermittent imagery fed into complex image analysis software, which is not especially accurate and is challenging to use effectively. We have developed a simple and fast alternative technique that uses a pinning robot and a commonplace microplate reader to continuously measure the thickness of colonies growing on solid agar, complemented by a technique for normalizing the amount of cells initially printed to each spot of the array in the first place. We have developed software to automate the process of combining multiple sets of readings, subtracting agar absorbance, and visualizing colony thickness changes in a number of informative ways.RESULTSThe “PHENOS” pipeline (PHENotyping On Solid media), optimized forSaccharomycesyeasts, produces highly reproducible growth curves and is particularly sensitive to low-level growth. We have empirically determined a formula to estimate colony cell count from an absorbance measurement, and shown this to be comparable with estimates from measurements in liquid. We have also validated the technique by reproducing the results of an earlier QTL study done with conventional liquid phenotyping, and found PHENOS to be considerably more sensitive.CONCLUSIONS“PHENOS” is a cost effective and reliable high-throughput technique for quantifying growth of yeast arrays, and is likely to be equally very useful for a range of other types of microbial arrays. A detailed guide to the pipeline and software is provided with the installation files athttps://github.com/gact/phenos.
Visual evidence accumulation guides decision-making in unrestrained mice
Authors: Odoemene Onyekachi , Pisupati Sashank , Nguyen Hien , Churchland Anne K. 
Published: September 2017

AbstractThe ability to manipulate neural activity with precision is an asset in uncovering neural circuits for decision-making. Diverse tools for manipulating neurons are available for mice, but the feasibility of mice for decision-making studies remains unclear, especially when decisions require accumulating visual evidence. For example, whether mice’ decisions reflect leaky accumulation is not established, and the relevant and irrelevant factors that influence decisions are unknown. Further, causal circuits for visual evidence accumulation have not been established. To address these issues, we measured >500,000 decisions in 27 mice trained to judge the fluctuating rate of a sequence of flashes. Information throughout the 1000ms trial influenced choice, but early information was most influential. This suggests that information persists in neural circuits for ~1000ms with minimal accumulation leak. Further, while animals primarily based decisions on current stimulus rate, they were unable to entirely suppress additional factors: total stimulus brightness and the previous trial’s outcome. Next, we optogenetically inhibited anteromedial (AM) visual area using JAWS. Importantly, light activation biased choices in both injected and uninjected animals, demonstrating that light alone influences behavior. By varying stimulus-response contingency while holding stimulated hemisphere constant, we surmounted this obstacle to demonstrate that AM suppression biases decisions. By leveraging a large dataset to quantitatively characterize decision-making behavior, we establish mice as suitable for neural circuit manipulation studies, including the one here. Further, by demonstrating that mice accumulate visual evidence, we demonstrate that this strategy for reducing uncertainty in decision-making is employed by animals with diverse visual systems.Significance statementTo connect behaviors to their underlying neural mechanism, a deep understanding of the behavioral strategy is needed. This understanding is incomplete in mouse studies, in part because existing datasets have been too small to quantitatively characterize decision-making behavior. To surmount this, we measured the outcome of over 500,000 decisions made by 27 mice trained to judge visual stimuli. Our analyses offer new insights into mice’ decision-making strategies and compares them with those of other species. We then disrupted neural activity in a candidate neural structure and examined the effect on decisions. Our findings establish mice as a suitable organism for visual accumulation of evidence decisions. Further, the results highlight similarities in decision-making strategies across very different species.
Simultaneous representation of a spectrum of dynamically changing value estimates during decision making
Authors: Meder David , Kolling Nils , Verhagen Lennart , Wittmann Marco K , Scholl Jacqueline , Madsen Kristoffer H , Hulme Oliver J , Behrens Timothy EJ , Rushworth Matthew FS 
Published: September 2017

SummaryDecisions are based on value expectations derived from experience. We show that dorsal anterior cingulate cortex and three other brain regions hold multiple representations of choice value based on different time-scales of experience organized in terms of systematic gradients across the cortex. Some parts of each area represent value estimates based on recent reward experience while others represent value estimates based on experience over the longer term. The value estimates within these four brain areas interact with one another according to their temporal scaling. Some aspects of the representations change dynamically as the environment changes. The spectrum of value estimates may act as a flexible selection mechanism for combining experience-derived value information with other aspects of value to allow flexible and adaptive decisions in changing environments.
Evaluation of MRI sequences for quantitative T1 brain mapping
Authors: Tsialios P , Thrippleton M , Pernet C 
Published: September 2017

AbstractT1 mapping constitutes a quantitative MRI technique finding significant application in brain imaging. It allows improved evaluation of contrast uptake, blood perfusion, volume, and provides more specific biomarkers of disease progression compared to conventional T1-weighted images. While there are many techniques for T1-mapping, there is also a wide range of reported T1-values in tissues, raising the issue of protocols’ reproducibility and standardization. The gold standard for obtaining T1-maps is based on acquiring IR-SE sequence. Widely used alternative sequences are IR-SE-EPI, VFA (DESPOT), DESPOT-HIFI and MP2RAGE that speed up scanning and fitting procedures. A custom MRI phantom was used to assess the reproducibility and accuracy of the different methods. All scans were performed using a 3T Siemens Prisma scanner. The acquired data were processed using two different codes. The main difference was observed for VFA (DESPOT) which grossly overestimated T1 relaxation time by 214 ms [CI: 126 270 ms] compared to the IR-SE sequence. MP2RAGE and DESPOT-HIFI sequences gave slightly shorter T1 than IR-SE (∼20 to 30ms) and can be considered as alternative and time-efficient methods for acquiring accurate T1 maps of the human brain, while IR-SE-EPI gave identical results, at a cost of a lower image quality.
Karrikin-sensing protein KAI2 is a new player in regulating root growth patterns
Authors: Swarbreck Stéphanie M. , Guerringue Yannick , Matthus Elsa , Jamieson Fiona J. C. , Davies Julia M. 
Published: September 2017

SummaryRoots form highly complex systems varying in growth direction and branching pattern to forage for nutrients efficiently. Here mutations in the KAI2 (KARRIKIN INSENSITIVE) α/β-fold hydrolase and the MAX2 (MORE AXILLARY GROWTH 2) F-box leucine-rich protein, which together perceive karrikins (smoke-derived butenolides), caused alteration in root growth direction (root skewing and waving) ofArabidopsis thaliana. This exaggerated root skewing was independent of endogenous strigolactone perception by the D14 α/β-fold hydrolase and MAX2. Thus KAI2/MAX2’s regulation of root growth may be through perception of endogenous KAI2-ligands, which have yet to be identified. Degradation targets of the KAI2/MAX2 complex, SMAX1 (SUPPRESSOR OF MAX2-1) and SMXL6,7,8 (SUPPRESSOR OF MAX2-1-LIKE) are also involved in the regulation of root skewing. Genetic data reveal a new potential target for degradation, as mutation in the SKS3 (SKU5 similar) but not the SKU5/SKS17 root plasma membrane glycoprotein suppresses the exaggerated root skewing induced by the lack of MAX2. InArabidopsis thalianatherefore, the KAI2 karrikin-sensing protein acts to limit root skewing, and we propose a mechanism involving root radial expansion as the mutant’s gravitropic and mechano-sensing responses remained largely unaffected.
Metabolic characterization of menopause: cross-sectional and longitudinal evidence
Authors: Wang Qin , Santos Ferreira Diana L , Nelson Scott M , Sattar Naveed , Ala-Korpela Mika , Lawlor Debbie A 
Published: September 2017

AbstractBackgroundIt remains elusive whether the changes in cardiometabolic biomarkers during the menopausal transition are due to ovarian aging or chronological aging. Well-conducted longitudinal studies are required to determine this. The aim of this study was to explore the cross-sectional and longitudinal associations of reproductive status defined according to the 2012 Stages of Reproductive Aging Workshop criteria with 74 metabolic biomarkers, and establish whether any associations are independent of age related changes.MethodsWe determined cross-sectional associations of reproductive status with metabolic profiling in 3,312 UK midlife women. In a subgroup of 1,492 women who had repeat assessments after 2.5 years, we assessed how change in reproductive status was associated with the changes in metabolic biomarkers. Metabolic profiles were measured by high-throughput quantitative serum NMR metabolomics. In longitudinal analyses, we compared the change in metabolic biomarkers for each reproductive status category change to that in the reference of being pre-menopausal at both time points. As all women aged by a similar amount during follow-up, these analyses contribute to distinguish age related changes from those related to change in reproductive status.ResultsConsistent cross-sectional and longitudinal associations of menopause with a wide range of metabolic biomarkers were observed, suggesting transition to menopause induces multiple metabolic changes independent of chronological aging. The metabolic changes included increased concentrations of very small VLDL, IDL and LDL subclasses, remnant and LDL cholesterol, and reduced LDL particle size, all towards an atherogenic lipoprotein profile. Increased inflammation was suggested via an inflammatory biomarker, glycoprotein acetyls, but not via C-reactive protein. Also, levels of glutamine and albumin were increased during the transition. Most of these metabolic changes seen at the time of becoming post-menopausal remained or became slightly stronger during the post-menopausal years.ConclusionsTransition to post-menopause has effects on multiple circulating metabolic biomarkers, over and above the underlying age trajectory. The adverse changes in multiple apolipoprotein-B containing lipoprotein subclasses and increased inflammation may underlie women’s increased cardiometabolic risk in post-menopausal years.AbbreviationsALSPACAvon Longitudinal Study of Parents and ChildrenBMIbody mass indexCRPhigh sensitive C-reactive proteinCVDcardiovascular diseasesHDLhigh-density lipoproteinHRThormone replacement therapyIDLintermediate-density lipoproteinLDLlow-density lipoproteinSDstandard deviationSTRAWStages of Reproductive Aging WorkshopSWANThe Study of Women’s health Across the NationVLDLvery low-density lipoprotein
Discovery of physiological and cancer-related regulators of 3’ UTR processing with KAPAC
Authors: Gruber Andreas J. , Schmidt Ralf , Ghosh Souvik , Martin Georges , Gruber Andreas R. , van Nimwegen Erik , Zavolan Mihaela 
Published: September 2017

Abstract3’ UTR length is regulated in relation to cellular state. To uncover key regulators of poly(A) site (PAS) use in specific conditions, we have developed PAQR, a method for quantifying PAS use from RNA sequencing data and KAPAC, an approach that infers activities of oligomeric sequence motifs on PAS choice. Application of PAQR and KAPAC to RNA sequencing data from normal and tumor tissue samples uncovered sequence motifs that can explain changes in cleavage and polyadenylation in specific cancers. In particular, our analysis points to Polypyrimidine tract binding protein 1 as a regulator of PAS choice in glioblastoma.
CRISPR-Based DNA Imaging in Living Cells Reveals Cell Cycle-Dependent Chromosome Dynamics
Authors: Ma Hanhui , Tu Li-Chun , Naseri Ardalan , Chung Yu-Chieh , Grunwald David , Zhang Shaojie , Pederson Thoru 
Published: September 2017

SUMMARYIn contrast to the well-studied condensation and folding of chromosomes during mitosis, their dynamics in interphase are less understood. We developed a sensitive, multicolor system, CRISPR-Sirius, allowing the real-time tracking of the dynamics of chromosomal loci. We tracked loci kilobases to megabases apart and found significant variation in the inter-locus distances of each pair, indicating differing degrees of DNA contortion. We resolved two distinct modes of dynamics of loci: saltatory local movements as well as translational movements of the domain. The magnitude of both of these modes of movements increased from early to late G1, whereas the translational movements were reduced in early S. The local fluctuations decreased slightly in early S and more markedly in mid-late S. These newly observed movements and their cell cycle-dependence are indicative of a hitherto unrecognized compaction-relaxation dynamic of the chromosomal fiber operating concurrently with changes in the extent of observed genomic domain movements.IN BRIEFDistinct chromosome folding and dynamics during cell cycle progression were dissected by CRISPR-Sirius DNA imaging in living cells.HIGHLIGHTSCRISPR-Sirius allows tracking of pairs of chromosomal loci having kilobase to megabase inter-locus distancesPair-wise tracking of loci allows measurement of both local and domain dynamicsChromosomal fiber relaxation is positively correlated with local dynamicsGenomic region size contributes to local and domain movementsDistinct chromosome dynamics were uncovered during cell cycle progression in interphase
State-Based Delay Representation and Its Transfer from a Game of Pong to Reaching and Tracking
Authors: Avraham Guy , Leib Raz , Pressman Assaf , Simo Lucia S. , Karniel Amir , Shmuelof Lior , Mussa-Ivaldi Ferdinando A. , Nisky Ilana 
Published: September 2017

AbstractTo accurately estimate the state of the body, the nervous system needs to account for delays between signals from different sensory modalities. To investigate how such delays may be represented in the sensorimotor system, we asked human participants to play a virtual pong game in which the movement of the virtual paddle was delayed with respect to their hand movement. We tested the representation of this new mapping between the hand and the delayed paddle by examining transfer of adaptation to blind reaching and blind tracking tasks. These blind tasks enabled to capture the representation in feedforward mechanisms of movement control. A Time Representation of the delay is an estimation of the actual time lag between hand and paddle movements. A State Representation is a representation of delay using current state variables: the distance between the paddle and the ball originating from the delay may be considered as a spatial shift; the low sensitivity in the response of the paddle may be interpreted as a minifying gain; and the lag may be attributed to a mechanical resistance that influences paddle’s movement. We found that the effects of prolonged exposure to the delayed feedback transferred to blind reaching and tracking tasks and caused participants to exhibit hypermetric movements. These results, together with simulations of our representation models, suggest that delay is not represented based on time, but rather as a spatial gain change in visuomotor mapping.Significance StatementIt is known that the brain copes with sensory feedback delays to control movements, but it is unclear whether it does so using a representation of the actual time lag. We addressed this question by exposing participants to a visuomotor delay during a dynamic game of pong. Following the game, participants exhibited hypermetric reaching and tracking movements that indicate that delay is represented as a visuomotor gain rather than as a temporal shift.
Embryonic and postnatal neurogenesis produce functionally distinct subclasses of dopaminergic neuron
Authors: Galliano Elisa , Franzoni Eleonora , Breton Marine , Chand Annisa N. , Byrne Darren J. , Murthy Venkatesh N. , Grubb Matthew S. 
Published: September 2017

AbstractMost neurogenesis in the mammalian brain is completed embryonically, but in certain areas the production of neurons continues throughout postnatal life. The functional properties of mature postnatally-generated neurons often match those of their embryonically-produced counterparts. However, we show here that in the olfactory bulb (OB), embryonic and postnatal neurogenesis produce functionally distinct subpopulations of dopaminergic (DA) neurons. We define two subclasses of OB DA neuron by the presence or absence of a key subcellular specialisation: the axon initial segment (AIS). Large AIS-positive axon-bearing DA neurons are exclusively produced during early embryonic stages, leaving small anaxonic AIS-negative cells as the only DA subtype generated via adult neurogenesis. These populations are functionally distinct: large DA cells are more excitable, yet display weaker and – for certain long-latency or inhibitory events – more broadly-tuned responses to odorant stimuli. Embryonic and postnatal neurogenesis can therefore generate distinct neuronal subclasses, placing important constraints on the functional roles of adult-born neurons in sensory processing.
Music training improves the ability to understand speech-in-noise in older adults
Authors: Zendel Benjamin Rich , West Greg , Belleville Sylvie , Peretz Isabelle 
Published: September 2017

AbstractIt is well known that hearing abilities decline with age, and one of the most commonly reported hearing difficulties reported in older adults is a reduced ability to understand speech in noisy environments. Older musicians have an enhanced ability to understand speech in noise, and this has been associated with enhanced brain responses related to both speech processing and the deployment of attention, however the causal impact of music lessons in older adults is poorly understood. A sample of older adults was randomly assigned to learn to play piano (Mus), to learn to play a visuo-spatially demanding video-game (Vid), or to serve as a no-contact control (Nocon).After 6 months, the Mus group improved their ability to understand a word presented in loud background noise. This improvement was related to an earlier N100, enhanced P250 (P2/P3) and a reduced N600 (N400). These findings support the idea that music lessons provide a causal benefit to hearing abilities, and that this benefit is due to both enhanced encoding of speech stimuli, and enhanced deployment of attentional mechanisms towards the speech stimuli. Importantly, these findings suggest that music training could be used as a foundation to develop auditory rehabilitation programs for older adults.