The assembly of DNA sequences de novo is fundamental to genomics research. It is the first of many steps toward elucidating and characterizing whole genomes. Downstream applications, including ...analysis of genomic variation between species, between or within individuals critically depend on robustly assembled sequences. In the span of a single decade, the sequence throughput of leading DNA sequencing instruments has increased drastically, and coupled with established and planned large-scale, personalized medicine initiatives to sequence genomes in the thousands and even millions, the development of efficient, scalable and accurate bioinformatics tools for producing high-quality reference draft genomes is timely. With ABySS 1.0, we originally showed that assembling the human genome using short 50-bp sequencing reads was possible by aggregating the half terabyte of compute memory needed over several computers using a standardized message-passing system (MPI). We present here its redesign, which departs from MPI and instead implements algorithms that employ a Bloom filter, a probabilistic data structure, to represent a de Bruijn graph and reduce memory requirements. We benchmarked ABySS 2.0 human genome assembly using a Genome in a Bottle data set of 250-bp Illumina paired-end and 6-kbp mate-pair libraries from a single individual. Our assembly yielded a NG50 (NGA50) scaffold contiguity of 3.5 (3.0) Mbp using <35 GB of RAM. This is a modest memory requirement by today's standards and is often available on a single computer. We also investigate the use of BioNano Genomics and 10x Genomics' Chromium data to further improve the scaffold NG50 (NGA50) of this assembly to 42 (15) Mbp.
We report on the synthesis of bivalent water‐soluble calix4arene and calix5arene hosts, Super‐sCx4 and Super‐sCx5 as new broad‐spectrum supramolecular binders of neuromuscular blocking agents ...(NMBAs). Synthesis was achieved using the target bisquaternary amine NMBAs as a template to link two highly anionic p‐sulfonatocalixarene building blocks in aqueous solution. Bivalent anionic hosts Super‐sCx4 and Super‐sCx5 bind by engaging both quaternary amines present on a variety of NMBAs. We report low μM binding to structurally diverse alkyl, steroidal, curarine and benzylisoquinoline NMBAs with high selectivity over the neurotransmitter acetylcholine and a variety of other hydrophobic amines.
Broad‐spectrum binding: New bivalent supramolecular reversal agents are formed by target templated synthesis in aqueous solution. Bivalent anionic Super‐sCx calixarene hosts form strong complexes to a variety of structurally diverse target neuromuscular blocking agents.
Generating high-quality de novo genome assemblies is foundational to the genomics study of model and non-model organisms. In recent years, long-read sequencing has greatly benefited genome assembly ...and scaffolding, a process by which assembled sequences are ordered and oriented through the use of long-range information. Long reads are better able to span repetitive genomic regions compared to short reads, and thus have tremendous utility for resolving problematic regions and helping generate more complete draft assemblies. Here, we present LongStitch, a scalable pipeline that corrects and scaffolds draft genome assemblies exclusively using long reads. LongStitch incorporates multiple tools developed by our group and runs in up to three stages, which includes initial assembly correction (Tigmint-long), followed by two incremental scaffolding stages (ntLink and ARKS-long). Tigmint-long and ARKS-long are misassembly correction and scaffolding utilities, respectively, previously developed for linked reads, that we adapted for long reads. Here, we describe the LongStitch pipeline and introduce our new long-read scaffolder, ntLink, which utilizes lightweight minimizer mappings to join contigs. LongStitch was tested on short and long-read assemblies of Caenorhabditis elegans, Oryza sativa, and three different human individuals using corresponding nanopore long-read data, and improves the contiguity of each assembly from 1.2-fold up to 304.6-fold (as measured by NGA50 length). Furthermore, LongStitch generates more contiguous and correct assemblies compared to state-of-the-art long-read scaffolder LRScaf in most tests, and consistently improves upon human assemblies in under five hours using less than 23 GB of RAM. Due to its effectiveness and efficiency in improving draft assemblies using long reads, we expect LongStitch to benefit a wide variety of de novo genome assembly projects. The LongStitch pipeline is freely available at https://github.com/bcgsc/longstitch.
Antibiotic resistance is a growing global health concern prompting researchers to seek alternatives to conventional antibiotics. Antimicrobial peptides (AMPs) are attracting attention again as ...therapeutic agents with promising utility in this domain, and using in silico methods to discover novel AMPs is a strategy that is gaining interest. Such methods can sift through large volumes of candidate sequences and reduce lab screening costs.
Here we introduce AMPlify, an attentive deep learning model for AMP prediction, and demonstrate its utility in prioritizing peptide sequences derived from the Rana Lithobates catesbeiana (bullfrog) genome. We tested the bioactivity of our predicted peptides against a panel of bacterial species, including representatives from the World Health Organization's priority pathogens list. Four of our novel AMPs were active against multiple species of bacteria, including a multi-drug resistant isolate of carbapenemase-producing Escherichia coli.
We demonstrate the utility of deep learning based tools like AMPlify in our fight against antibiotic resistance. We expect such tools to play a significant role in discovering novel candidates of peptide-based alternatives to classical antibiotics.
Tandem repeat (TR) expansion is the underlying cause of over 40 neurological disorders. Long-read sequencing offers an exciting avenue over conventional technologies for detecting TR expansions. ...Here, we present Straglr, a robust software tool for both targeted genotyping and novel expansion detection from long-read alignments. We benchmark Straglr using various simulations, targeted genotyping data of cell lines carrying expansions of known diseases, and whole genome sequencing data with chromosome-scale assembly. Our results suggest that Straglr may be useful for investigating disease-associated TR expansions using long-read sequencing.
Genome sequencing yields the sequence of many short snippets of DNA (reads) from a genome. Genome assembly attempts to reconstruct the original genome from which these reads were derived. This task ...is difficult due to gaps and errors in the sequencing data, repetitive sequence in the underlying genome, and heterozygosity. As a result, assembly errors are common. In the absence of a reference genome, these misassemblies may be identified by comparing the sequencing data to the assembly and looking for discrepancies between the two. Once identified, these misassemblies may be corrected, improving the quality of the assembled sequence. Although tools exist to identify and correct misassemblies using Illumina paired-end and mate-pair sequencing, no such tool yet exists that makes use of the long distance information of the large molecules provided by linked reads, such as those offered by the 10x Genomics Chromium platform. We have developed the tool Tigmint to address this gap.
To demonstrate the effectiveness of Tigmint, we applied it to assemblies of a human genome using short reads assembled with ABySS 2.0 and other assemblers. Tigmint reduced the number of misassemblies identified by QUAST in the ABySS assembly by 216 (27%). While scaffolding with ARCS alone more than doubled the scaffold NGA50 of the assembly from 3 to 8 Mbp, the combination of Tigmint and ARCS improved the scaffold NGA50 of the assembly over five-fold to 16.4 Mbp. This notable improvement in contiguity highlights the utility of assembly correction in refining assemblies. We demonstrate the utility of Tigmint in correcting the assemblies of multiple tools, as well as in using Chromium reads to correct and scaffold assemblies of long single-molecule sequencing.
Scaffolding an assembly that has been corrected with Tigmint yields a final assembly that is both more correct and substantially more contiguous than an assembly that has not been corrected. Using single-molecule sequencing in combination with linked reads enables a genome sequence assembly that achieves both a high sequence contiguity as well as high scaffold contiguity, a feat not currently achievable with either technology alone.
The comprehensive multiplatform genomics data generated by The Cancer Genome Atlas (TCGA) Research Network is an enabling resource for cancer research. It includes an unprecedented amount of microRNA ...sequence data: ~11 000 libraries across 33 cancer types. Combined with initiatives like the National Cancer Institute Genomics Cloud Pilots, such data resources will make intensive analysis of large-scale cancer genomics data widely accessible. To support such initiatives, and to enable comparison of TCGA microRNA data to data from other projects, we describe the process that we developed and used to generate the microRNA sequence data, from library construction through to submission of data to repositories. In the context of this process, we describe the computational pipeline that we used to characterize microRNA expression across large patient cohorts.
Antibiotic resistance is recognized as an imminent and growing global health threat. New antimicrobial drugs are urgently needed due to the decreasing effectiveness of conventional small‐molecule ...antibiotics. Antimicrobial peptides (AMPs), a class of host defense peptides, are emerging as promising candidates to address this need. The potential sequence space of amino acids is combinatorially vast, making it possible to extend the current arsenal of antimicrobial agents with a practically infinite number of new peptide‐based candidates. However, mining naturally occurring AMPs, whether directly by wet lab screening methods or aided by bioinformatics prediction tools, has its theoretical limit regarding the number of samples or genomic/transcriptomic resources researchers have access to. Further, manually designing novel synthetic AMPs requires prior field knowledge, restricting its throughput. In silico sequence generation methods are gaining interest as a high‐throughput solution to the problem. Here, we introduce AMPd‐Up, a recurrent neural network based tool for de novo AMP design, and demonstrate its utility over existing methods. Validation of candidates designed by AMPd‐Up through antimicrobial susceptibility testing revealed that 40 of the 58 generated sequences possessed antimicrobial activity against Escherichia coli and/or Staphylococcus aureus. These results illustrate that AMPd‐Up can be used to design novel synthetic AMPs with potent activities.
Antimicrobial resistance is a critical public health concern, necessitating the exploration of alternative treatments. While antimicrobial peptides (AMPs) show promise, assessing their toxicity using ...traditional wet lab methods is both time‐consuming and costly. We introduce tAMPer, a novel multi‐modal deep learning model designed to predict peptide toxicity by integrating the underlying amino acid sequence composition and the three‐dimensional structure of peptides. tAMPer adopts a graph‐based representation for peptides, encoding ColabFold‐predicted structures, where nodes represent amino acids and edges represent spatial interactions. Structural features are extracted using graph neural networks, and recurrent neural networks capture sequential dependencies. tAMPer's performance was assessed on a publicly available protein toxicity benchmark and an AMP hemolysis data we generated. On the latter, tAMPer achieves an F1‐score of 68.7%, outperforming the second‐best method by 23.4%. On the protein benchmark, tAMPer exhibited an improvement of over 3.0% in the F1‐score compared to current state‐of‐the‐art methods. We anticipate tAMPer to accelerate AMP discovery and development by reducing the reliance on laborious toxicity screening experiments.