Project description:We identified hankyphage prophages within B. thetaiotaomicron isolates gathered from French hospitals. We extracted genomic DNA from an overnight culture from a single colony of each strain and sequenced them using Nanopore sequencing using the Plasmidsaurus platform. This long-read approach helped the assembly of the phages and determination of the hankyphage ends. We also improved the annotation of the reference hankyphage (hankyphage p00 from P. dorei HM719) using a structural prediction approach and annotated our B. thetaiotaomicron hankyphages using this new annotation. In this project we upload the genomic raw reads of nanopore sequencing of our hankyphage-bearing B. thetaiotaomicron collection (jmh strains) and the processed assembled hankyphages.
Project description:Analyses of ancient DNA typically involve sequencing the surviving short oligonucleotides, and aligning to genome assemblies from related, modern species. Here, we report that skin from a female woolly mammoth (†Mammuthus primigenius) which died 52,000 years ago retained its ancient genome architecture. We use PaleoHi-C to map chromatin contacts and assemble its genome, yielding twenty-eight chromosome-length scaffolds. Chromosome territories, compartments, loops, Barr bodies, and inactive-X chromosome (Xi) superdomains persist. The active and inactive genome compartments in mammoth skin more closely resemble Asian elephant skin than other elephant tissues. Our analyses uncover new biology. Differences in compartmentalization reveal genes whose transcription was potentially altered in mammoths vs. elephants. Mammoth Xi has a tetradic architecture, not bipartite like human and mouse. We hypothesize that, shortly after this mammoth’s death, the sample spontaneously freeze-dried in the Siberian cold, leading to a glass transition that preserved subfossils of ancient chromosomes at nanometer scale.
Project description:Transfer RNAs are the fundamental adapter molecules of protein synthesis and the most abundant and heterogeneous class of noncoding RNA molecules in cells. The study of tRNA repertoires remains challenging, complicated by the presence of dozens of post transcriptional modifications. Nanopore sequencing is an emerging technology with promise for both tRNA sequencing and the detection of RNA modifications; however, such studies have been limited by the throughput and accuracy of direct RNA sequencing methods. Moreover, detection of the complete set of tRNA modifications by nanopore sequencing remains challenging. Here we show that recent updates to nanopore direct RNA sequencing chemistry (RNA004) combined with our own optimizations to tRNA sequencing protocols and analysis workflows enable high throughput coverage of tRNA molecules and characterization of nanopore signals produced by 43 distinct RNA modifications. We share best practices and protocols for nanopore sequencing of tRNA and further report successful detection of low abundance mitochondrial and viral tRNAs, providing proof of concept for use of nanopore sequencing to study tRNA populations in the context of infection and organelle biology. This work provides a roadmap to guide future efforts towards de novo detection of RNA modifications across multiple organisms using nanopore sequencing.
Project description:5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) are modified versions of cytosine in DNA with roles in regulating gene expression. Using whole genomic DNA from mouse cerebellum, we benchmark 5mC and 5hmC detection by Oxford Nanopore Technologies sequencing against other standard techniques. In addition, we assess the ability of duplex base-calling to study strand asymmetric modification. Nanopore detection of 5mC and 5hmC is accurate relative to compared techniques and opens new means of studying these modifications. Strand asymmetric modification is widespread across the genome but reduced at imprinting control regions and CTCF binding sites in mouse cerebellum. Here we demonstrate the unique ability of nanopore sequencing to improve the resolution and detail of cytosine modification mapping.
Project description:We used the nanopore Cas9 targeted sequencing (nCATS) strategy to specifically sequence 125 L1HS-containing loci in parallel and measure their DNA methylation levels using nanopore long-read sequencing. Each targeted locus is sequenced at high coverage (~45X) with unambiguously mapped reads spanning the entire L1 element, as well as its flanking sequences over several kilobases. The genome-wide profile of L1 methylation was also assessed by bs-ATLAS-seq in the same cell lines (E-MTAB-10895).
Project description:Nanopore sequencing data of rolling-circle amplification (RCA) products performed on hES cells, treated with 2mJ UV-C to induce apoptosis. The experimental conditions were designed to characterize circular DNA forms generated during apoptosis. gDNA from untreated and UV-treated cells are used as controls.
Project description:Oxford Nanopore enables direct RNA sequencing allowing for base calling of RNA modifications. We tested mouse hippocampi RNA samples, using a Nanopore direct RNA-seq protocol that in addition to long poly A selected RNAs allows sequencing also of non-poly A RNAs as well as short RNAs < 200nt (including SINE B2 RNAs and other non poly A non coding RNAs). We provide here as a resource a direct RNA-sequencing dataset generated from mouse brain tissues that includes both mRNAs and non poly A or short non-coding RNAs such as SINEs. Elevated SINE B2 RNA Adenosine to Inosine editing is consistently observed across hippocampal tissues of a mouse model of amyloid beta accumulation compared to hippocampi of wild type animals. Nanopore direct RNA sequencing supports increased RNA modification signals at the sameSINE B2 RNA regions identified by short-read Illumina sequencing in these hippocampi.