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  1. Abstract

    Sequencing, assembly, and annotation of the 26.5 Gbp hexaploid genome of coast redwood (Sequoia sempervirens) was completed leading toward discovery of genes related to climate adaptation and investigation of the origin of the hexaploid genome. Deep-coverage short-read Illumina sequencing data from haploid tissue from a single seed were combined with long-read Oxford Nanopore Technologies sequencing data from diploid needle tissue to create an initial assembly, which was then scaffolded using proximity ligation data to produce a highly contiguous final assembly, SESE 2.1, with a scaffold N50 size of 44.9 Mbp. The assembly included several scaffolds that span entire chromosome arms, confirmed by the presence of telomere and centromere sequences on the ends of the scaffolds. The structural annotation produced 118,906 genes with 113 containing introns that exceed 500 Kbp in length and one reaching 2 Mb. Nearly 19 Gbp of the genome represented repetitive content with the vast majority characterized as long terminal repeats, with a 2.9:1 ratio of Copia to Gypsy elements that may aid in gene expression control. Comparison of coast redwood to other conifers revealed species-specific expansions for a plethora of abiotic and biotic stress response genes, including those involved in fungal disease resistance, detoxification, and physical injury/structural remodeling and othersmore »supporting flavonoid biosynthesis. Analysis of multiple genes that exist in triplicate in coast redwood but only once in its diploid relative, giant sequoia, supports a previous hypothesis that the hexaploidy is the result of autopolyploidy rather than any hybridizations with separate but closely related conifer species.

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  2. Shao, Mingfu (Ed.)
    Third-generation sequencing technologies can generate very long reads with relatively high error rates. The lengths of the reads, which sometimes exceed one million bases, make them invaluable for resolving complex repeats that cannot be assembled using shorter reads. Many high-quality genome assemblies have already been produced, curated, and annotated using the previous generation of sequencing data, and full re-assembly of these genomes with long reads is not always practical or cost-effective. One strategy to upgrade existing assemblies is to generate additional coverage using long-read data, and add that to the previously assembled contigs. SAMBA is a tool that is designed to scaffold and gap-fill existing genome assemblies with additional long-read data, resulting in substantially greater contiguity. SAMBA is the only tool of its kind that also computes and fills in the sequence for all spanned gaps in the scaffolds, yielding much longer contigs. Here we compare SAMBA to several similar tools capable of re-scaffolding assemblies using long-read data, and we show that SAMBA yields better contiguity and introduces fewer errors than competing methods. SAMBA is open-source software that is distributed at .
    Free, publicly-accessible full text available February 4, 2023
  3. Valencia, Alfonso (Ed.)
    Abstract Motivation Improvements in DNA sequencing technology and computational methods have led to a substantial increase in the creation of high-quality genome assemblies of many species. To understand the biology of these genomes, annotation of gene features and other functional elements is essential; however, for most species, only the reference genome is well-annotated. Results One strategy to annotate new or improved genome assemblies is to map or ‘lift over’ the genes from a previously annotated reference genome. Here, we describe Liftoff, a new genome annotation lift-over tool capable of mapping genes between two assemblies of the same or closely related species. Liftoff aligns genes from a reference genome to a target genome and finds the mapping that maximizes sequence identity while preserving the structure of each exon, transcript and gene. We show that Liftoff can accurately map 99.9% of genes between two versions of the human reference genome with an average sequence identity >99.9%. We also show that Liftoff can map genes across species by successfully lifting over 98.3% of human protein-coding genes to a chimpanzee genome assembly with 98.2% sequence identity. Availability and implementation Liftoff can be installed via bioconda and PyPI. In addition, the source code for Liftoffmore »is available at Supplementary information Supplementary data are available at Bioinformatics online.« less
  4. Understanding the genomic and environmental basis of cold adaptation is key to understand how plants survive and adapt to different environmental conditions across their natural range. Univariate and multivariate genome-wide association (GWAS) and genotype-environment association (GEA) analyses were used to test associations among genome-wide SNPs obtained from whole-genome resequencing, measures of growth, phenology, emergence, cold hardiness, and range-wide environmental variation in coastal Douglas-fir (Pseudotsuga menziesii). Results suggest a complex genomic architecture of cold adaptation, in which traits are either highly polygenic or controlled by both large and small effect genes. Newly discovered associations for cold adaptation in Douglas-fir included 130 genes involved in many important biological functions such as primary and secondary metabolism, growth and reproductive development, transcription regulation, stress and signaling, and DNA processes. These genes were related to growth, phenology and cold hardiness and strongly depend on variation in environmental variables such degree days below 0c, precipitation, elevation and distance from the coast. This study is a step forward in our understanding of the complex interconnection between environment and genomics and their role in cold-associated trait variation in boreal tree species, providing a baseline for the species’ predictions under climate change.
  5. Rzhetsky, Andrey (Ed.)
    GC skew is a phenomenon observed in many bacterial genomes, wherein the two replication strands of the same chromosome contain different proportions of guanine and cytosine nucleotides. Here we demonstrate that this phenomenon, which was first discovered in the mid-1990s, can be used today as an analysis tool for the 15,000+ complete bacterial genomes in NCBI’s Refseq library. In order to analyze all 15,000+ genomes, we introduce a new method, SkewIT (Skew Index Test), that calculates a single metric representing the degree of GC skew for a genome. Using this metric, we demonstrate how GC skew patterns are conserved within certain bacterial phyla, e.g. Firmicutes, but show different patterns in other phylogenetic groups such as Actinobacteria. We also discovered that outlier values of SkewIT highlight potential bacterial mis-assemblies. Using our newly defined metric, we identify multiple mis-assembled chromosomal sequences in previously published complete bacterial genomes. We provide a SkewIT web app that calculates SkewI for any user-provided bacterial sequence. The web app also provides an interactive interface for the data generated in this paper, allowing users to further investigate the SkewI values and thresholds of the Refseq-97 complete bacterial genomes. Individual scripts for analysis of bacterial genomes are providedmore »in the following repository: .« less
  6. Abstract The giant sequoia (Sequoiadendron giganteum) of California are massive, long-lived trees that grow along the U.S. Sierra Nevada mountains. Genomic data are limited in giant sequoia and producing a reference genome sequence has been an important goal to allow marker development for restoration and management. Using deep-coverage Illumina and Oxford Nanopore sequencing, combined with Dovetail chromosome conformation capture libraries, the genome was assembled into eleven chromosome-scale scaffolds containing 8.125 Gbp of sequence. Iso-Seq transcripts, assembled from three distinct tissues, was used as evidence to annotate a total of 41,632 protein-coding genes. The genome was found to contain, distributed unevenly across all 11 chromosomes and in 63 orthogroups, over 900 complete or partial predicted NLR genes, of which 375 are supported by annotation derived from protein evidence and gene modeling. This giant sequoia reference genome sequence represents the first genome sequenced in the Cupressaceae family, and lays a foundation for using genomic tools to aid in giant sequoia conservation and management.
  7. Bread wheat (Triticum aestivum) is a major food crop and an important plant system for agricultural genetics research. However, due to the complexity and size of its allohexaploid genome, genomic resources are limited compared to other major crops. The IWGSC recently published a reference genome and associated annotation (IWGSC CS v1.0, Chinese Spring) that has been widely adopted and utilized by the wheat community. Although this reference assembly represents all three wheat subgenomes at chromosome-scale, it was derived from short reads, and thus is missing a substantial portion of the expected 16 Gbp of genomic sequence. We earlier published an independent wheat assembly (Triticum_aestivum_3.1, Chinese Spring) that came much closer in length to the expected genome size, although it was only a contig-level assembly lacking gene annotations. Here, we describe a reference-guided effort to scaffold those contigs into chromosome-length pseudomolecules, add in any missing sequence that was unique to the IWGSC CS v1.0 assembly, and annotate the resulting pseudomolecules with genes. Our updated assembly, Triticum_aestivum_4.0, contains 15.07 Gbp of non-gap sequence anchored to chromosomes, which is 1.2 Gbps more than the previous reference assembly. It includes 108,639 genes unambiguously localized to chromosomes, including over 2,000 genes that were previously unplaced.more »We also discovered more than 5,700 additional gene copies, facilitating the accurate annotation of functional gene duplications including at the Ppd-B1 photoperiod response locus.« less