Chromatin accessibility dynamics of myogenesis at single cell resolution, bioRxiv, 2017-06-27
AbstractOver a million DNA regulatory elements have been cataloged in the human genome, but linking these elements to the genes that they regulate remains challenging. We introduce Cicero, a statistical method that connects regulatory elements to target genes using single cell chromatin accessibility data. We apply Cicero to investigate how thousands of dynamically accessible elements orchestrate gene regulation in differentiating myoblasts. Groups of co-accessible regulatory elements linked by Cicero meet criteria of “chromatin hubs”, in that they are physically proximal, interact with a common set of transcription factors, and undergo coordinated changes in histone marks that are predictive of gene expression. Pseudotemporal analysis revealed a subset of elements bound by MYOD in myoblasts that exhibit early opening, potentially serving as the initial sites of recruitment of chromatin remodeling and histone-modifying enzymes. The methodological framework described here constitutes a powerful new approach for elucidating the architecture, grammar and mechanisms of cis-regulation on a genome-wide basis.
biorxiv genomics 100-200-users 2017Identification of a novel interspecific hybrid yeast from a metagenomic spontaneously inoculated beer sample using Hi-C, bioRxiv, 2017-06-16
AbstractInterspecific hybridization is a common mechanism enabling genetic diversification and adaptation; however, the detection of hybrid species has been quite difficult. The identification of microbial hybrids is made even more complicated, as most environmental microbes are resistant to culturing and must be studied in their native mixed communities. We have previously adapted the chromosome conformation capture method Hi-C to the assembly of genomes from mixed populations. Here, we show the method’s application in assembling genomes directly from an uncultured, mixed population from a spontaneously inoculated beer sample. Our assembly method has enabled us to de-convolute 4 bacterial and 4 yeast genomes from this sample, including a putative yeast hybrid. Downstream isolation and analysis of this hybrid confirmed its genome to consist of Pichia membranifaciens and that of another related, but undescribed yeast. Our work shows that Hi-C-based metagenomic methods can overcome the limitation of traditional sequencing methods in studying complex mixtures of genomes.
biorxiv genomics 0-100-users 2017High contiguity Arabidopsis thaliana genome assembly with a single nanopore flow cell, bioRxiv, 2017-06-15
AbstractWhile many evolutionary questions can be answered by short read re-sequencing, presenceabsence polymorphisms of genes andor transposons have been largely ignored in large-scale intraspecific evolutionary studies. To enable the rigorous analysis of such variants, multiple high quality and contiguous genome assemblies are essential. Similarly, while genome assemblies based on short reads have made genomics accessible for non-reference species, these assemblies have limitations due to low contiguity. Long-read sequencers and long-read technologies have ushered in a new era of genome sequencing where the lengths of reads exceed those of most repeats. However, because these technologies are not only costly, but also time and compute intensive, it has been unclear how scalable they are. Here we demonstrate a fast and cost effective reference assembly for an Arabidopsis thaliana accession using the USB-sized Oxford Nanopore MinION sequencer and typical consumer computing hardware (4 Cores, 16Gb RAM). We assemble the accession KBS-Mac-74 into 62 contigs with an N50 length of 12.3 Mb covering 100% (119 Mb) of the non-repetitive genome. We demonstrate that the polished KBS-Mac-74 assembly is highly contiguous with BioNano optical genome maps, and of high per-base quality against a likewise polished Pacific Biosciences long-read assembly. The approach we implemented took a total of four days at a cost of less than 1,000 USD for sequencing consumables including instrument depreciation.
biorxiv genomics 200-500-users 2017Regional missense constraint improves variant deleteriousness prediction, bioRxiv, 2017-06-13
AbstractGiven increasing numbers of patients who are undergoing exome or genome sequencing, it is critical to establish tools and methods to interpret the impact of genetic variation. While the ability to predict deleteriousness for any given variant is limited, missense variants remain a particularly challenging class of variation to interpret, since they can have drastically different effects depending on both the precise location and specific amino acid substitution of the variant. In order to better evaluate missense variation, we leveraged the exome sequencing data of 60,706 individuals from the Exome Aggregation Consortium (ExAC) dataset to identify sub-genic regions that are depleted of missense variation. We further used this depletion as part of a novel missense deleteriousness metric named MPC. We applied MPC to de novo missense variants and identified a category of de novo missense variants with the same impact on neurodevelopmental disorders as truncating mutations in intolerant genes, supporting the value of incorporating regional missense constraint in variant interpretation.
biorxiv genomics 100-200-users 2017Indexcov fast coverage quality control for whole-genome sequencing, bioRxiv, 2017-06-10
AbstractThe BAM1 and CRAM2 formats provide a supplementary linear index that facilitates rapid access to sequence alignments in arbitrary genomic regions. Comparing consecutive entries in a BAM or CRAM index allows one to infer the number of alignment records per genomic region for use as an effective proxy of sequence depth in each genomic region. Based on these properties, we have developed indexcov, an efficient estimator of whole-genome sequencing coverage to rapidly identify samples with aberrant coverage profiles, reveal large scale chromosomal anomalies, recognize potential batch effects, and infer the sex of a sample. Indexcov is available at <jatsext-link xmlnsxlink=httpwww.w3.org1999xlink ext-link-type=uri xlinkhref=httphttpsgithub.combrentpgoleft>httpsgithub.combrentpgoleft<jatsext-link> under the MIT license.
biorxiv genomics 100-200-users 2017Highly parallel genome variant engineering with CRISPRCas9 in eukaryotic cells, bioRxiv, 2017-06-09
AbstractDirect measurement of functional effects of DNA sequence variants throughout a genome is a major challenge. We developed a method that uses CRISPRCas9 to engineer many specific variants of interest in parallel in the budding yeast Saccharomyces cerevisiae, and to screen them for functional effects. We used the method to examine the functional consequences of premature termination codons (PTCs) at different locations within all annotated essential genes in yeast. We found that most PTCs were highly deleterious unless they occurred close to the C-terminal end and did not interrupt an annotated protein domain. Surprisingly, we discovered that some putatively essential genes are dispensable, while others have large dispensable regions. This approach can be used to profile the effects of large classes of variants in a high-throughput manner.
biorxiv genomics 0-100-users 2017