Direct visualization of transcriptional activation by physical enhancer–promoter proximity, bioRxiv, 2017-01-12
A long-standing question in metazoan gene regulation is how remote enhancers communicate with their target promoters over long distances. Combining genome editing and quantitative live imaging we simultaneously visualize physical enhancer–promoter communication and transcription in Drosophila embryos. Enhancers regulating pair rule stripes of even-skipped expression activate transcription of a reporter gene over a distance of 150 kb. We show in individual cells that activation only occurs after the enhancer comes into close proximity with its regulatory target and that upon dissociation transcription ceases almost immediately. We further observe distinct topological conformations of the eve locus, depending on the spatial identity of the activating stripe enhancer. In addition, long-range activation results in transcriptional competition at the endogenous eve locus, causing corresponding developmental defects. Overall, we demonstrate that sustained physical proximity and enhancer–promoter engagement are required for enhancer action, and we provide a path to probe the implications of long-range regulation on cellular fates.
biorxiv biophysics 100-200-users 2017Critical Assessment of Metagenome Interpretation – a benchmark of computational metagenomics software, bioRxiv, 2017-01-10
AbstractIn metagenome analysis, computational methods for assembly, taxonomic profiling and binning are key components facilitating downstream biological data interpretation. However, a lack of consensus about benchmarking datasets and evaluation metrics complicates proper performance assessment. The Critical Assessment of Metagenome Interpretation (CAMI) challenge has engaged the global developer community to benchmark their programs on datasets of unprecedented complexity and realism. Benchmark metagenomes were generated from ~700 newly sequenced microorganisms and ~600 novel viruses and plasmids, including genomes with varying degrees of relatedness to each other and to publicly available ones and representing common experimental setups. Across all datasets, assembly and genome binning programs performed well for species represented by individual genomes, while performance was substantially affected by the presence of related strains. Taxonomic profiling and binning programs were proficient at high taxonomic ranks, with a notable performance decrease below the family level. Parameter settings substantially impacted performances, underscoring the importance of program reproducibility. While highlighting current challenges in computational metagenomics, the CAMI results provide a roadmap for software selection to answer specific research questions.
biorxiv bioinformatics 100-200-users 2017Multiplex PCR method for MinION and Illumina sequencing of Zika and other virus genomes directly from clinical samples, bioRxiv, 2017-01-10
Genome sequencing has become a powerful tool for studying emerging infectious diseases; however, genome sequencing directly from clinical samples without isolation remains challenging for viruses such as Zika, where metagenomic sequencing methods may generate insufficient numbers of viral reads. Here we present a protocol for generating coding-sequence complete genomes comprising an online primer design tool, a novel multiplex PCR enrichment protocol, optimised library preparation methods for the portable MinION sequencer (Oxford Nanopore Technologies) and the Illumina range of instruments, and a bioinformatics pipeline for generating consensus sequences. The MinION protocol does not require an internet connection for analysis, making it suitable for field applications with limited connectivity. Our method relies on multiplex PCR for targeted enrichment of viral genomes from samples containing as few as 50 genome copies per reaction. Viral consensus sequences can be achieved starting with clinical samples in 1-2 days following a simple laboratory workflow. This method has been successfully used by several groups studying Zika virus evolution and is facilitating an understanding of the spread of the virus in the Americas.
biorxiv genomics 100-200-users 2017Whole genome sequencing and assembly of a Caenorhabditis elegans genome with complex genomic rearrangements using the MinION sequencing device, bioRxiv, 2017-01-09
ABSTRACTAdvances in 3rd generation sequencing have opened new possibilities for ‘benchtop’ whole genome sequencing. The MinION is a portable device that uses nanopore technology and can sequence long DNA molecules. MinION long reads are well suited for sequencing and de novo assembly of complex genomes with large repetitive elements. Long reads also facilitate the identification of complex genomic rearrangements such as those observed in tumor genomes. To assess the feasibility of the de novo assembly of large complex genomes using both MinION and Illumina platforms, we sequenced the genome of a Caenorhabditis elegans strain that contains a complex acetaldehyde-induced rearrangement and a biolistic bombardment-mediated insertion of a GFP containing plasmid. Using ∼5.8 gigabases of MinION sequence data, we were able to assemble a C. elegans genome containing 145 contigs (N50 contig length = 1.22 Mb) that covered >99% of the 100,286,401 bp reference genome. In contrast, using ∼8.04 gigabases of Illumina sequence data, we were able to assemble a C. elegans genome in 38,645 contigs (N50 contig length = ∼26 kb) containing 117 Mb. From the MinION genome assembly we identified the complex structures of both the acetaldehyde-induced mutation and the biolistic-mediated insertion. To date, this is the largest genome to be assembled exclusively from MinION data and is the first demonstration that the long reads of MinION sequencing can be used for whole genome assembly of large (100 Mb) genomes and the elucidation of complex genomic rearrangements.
biorxiv genomics 100-200-users 2017Neural precursors of deliberate and arbitrary decisions in the study of voluntary action, bioRxiv, 2017-01-02
AbstractThe readiness potential (RP)—a key ERP correlate of upcoming action—is known to precede subjects’ reports of their decision to move. Some view this as evidence against a causal role for consciousness in human decision-making and thus against free-will. Yet those studies focused on arbitrary decisions—purposeless, unreasoned, and without consequences. It remains unknown to what degree the RP generalizes to deliberate, more ecological decisions. We directly compared deliberate and arbitrary decision-making during a $1000-donation task to non-profit organizations. While we found the expected RPs for arbitrary decisions, they were strikingly absent for deliberate ones. Our results and drift-diffusion model are congruent with the RP representing accumulation of noisy, random fluctuations that drive arbitrary—but not deliberate—decisions. They further point to different neural mechanisms underlying deliberate and arbitrary decisions, challenging the generalizability of studies that argue for no causal role for consciousness in decision-making to real-life decisions.Significance StatementThe extent of human free will has been debated for millennia. Previous studies demonstrated that neural precursors of action—especially the readiness potential—precede subjects’ reports of deciding to move. Some viewed this as evidence against free-will. However, these experiments focused on arbitrary decisions—e.g., randomly raising the left or right hand. We directly compared deliberate (actual $1000 donations to NPOs) and arbitrary decisions, and found readiness potentials before arbitrary decisions, but—critically—not before deliberate decisions. This supports the interpretation of readiness potentials as byproducts of accumulation of random fluctuations in arbitrary but not deliberate decisions and points to different neural mechanisms underlying deliberate and arbitrary choice. Hence, it challenges the generalizability of previous results from arbitrary to deliberate decisions.
biorxiv neuroscience 100-200-users 2017A Data Citation Roadmap for Scholarly Data Repositories, bioRxiv, 2016-12-29
AbstractThis article presents a practical roadmap for scholarly data repositories to implement data citation in accordance with the Joint Declaration of Data Citation Principles, a synopsis and harmonization of the recommendations of major science policy bodies. The roadmap was developed by the Repositories Expert Group, as part of the Data Citation Implementation Pilot (DCIP) project, an initiative of FORCE11.org and the NIH BioCADDIE (<jatsext-link xmlnsxlink=httpwww.w3.org1999xlink ext-link-type=uri xlinkhref=httpsbiocaddie.org>httpsbiocaddie.org<jatsext-link>) program. The roadmap makes 11 specific recommendations, grouped into three phases of implementation a) required steps needed to support the Joint Declaration of Data Citation Principles, b) recommended steps that facilitate articledata publication workflows, and c) optional steps that further improve data citation support provided by data repositories.
biorxiv scientific-communication-and-education 200-500-users 2016