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Asset Tag: Large genome

April 21, 2020

Neopinone isomerase is involved in codeine and morphine biosynthesis in opium poppy.

The isomerization of neopinone to codeinone is a critical step in the biosynthesis of opiate alkaloids in opium poppy. Previously assumed to be spontaneous, the process is in fact catalyzed enzymatically by neopinone isomerase (NISO). Without NISO the primary metabolic products in the plant, in engineered microbes and in vitro are neopine and neomorphine, which are structural isomers of codeine and morphine, respectively. Inclusion of NISO in yeast strains engineered to convert thebaine to natural or semisynthetic opiates dramatically enhances formation of the desired products at the expense of neopine and neomorphine accumulation. Along with thebaine synthase, NISO is the second member of the pathogenesis-related 10 (PR10) protein family recently implicated in the enzymatic catalysis of a presumed spontaneous conversion in morphine biosynthesis.

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April 21, 2020

Phased genome sequence of an interspecific hybrid flowering cherry, ‘Somei-Yoshino’ (Cerasus × yedoensis).

We report the phased genome sequence of an interspecific hybrid, the flowering cherry ‘Somei-Yoshino’ (Cerasus × yedoensis). The sequence data were obtained by single-molecule real-time sequencing technology, split into two subsets based on genome information of the two probable ancestors, and assembled to obtain two haplotype phased genome sequences of the interspecific hybrid. The resultant genome assembly consisting of the two haplotype sequences spanned 690.1 Mb with 4,552 contigs and an N50 length of 1.0 Mb. We predicted 95,076 high-confidence genes, including 94.9% of the core eukaryotic genes. Based on a high-density genetic map, we established a pair of eight pseudomolecule sequences, with highly conserved structures between the two haplotype sequences with 2.4 million sequence variants. A whole genome resequencing analysis of flowering cherries suggested that ‘Somei-Yoshino’ might be derived from a cross between C. spachiana and either C. speciosa or its relatives. A time-course transcriptome analysis of floral buds and flowers suggested comprehensive changes in gene expression in floral bud development towards flowering. These genome and transcriptome data are expected to provide insights into the evolution and cultivation of flowering cherry and the molecular mechanism underlying flowering. © The Author(s) 2019. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.

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April 21, 2020

Genome assembly and gene expression in the American black bear provides new insights into the renal response to hibernation.

The prevalence of chronic kidney disease (CKD) is rising worldwide and 10-15% of the global population currently suffers from CKD and its complications. Given the increasing prevalence of CKD there is an urgent need to find novel treatment options. The American black bear (Ursus americanus) copes with months of lowered kidney function and metabolism during hibernation without the devastating effects on metabolism and other consequences observed in humans. In a biomimetic approach to better understand kidney adaptations and physiology in hibernating black bears, we established a high-quality genome assembly. Subsequent RNA-Seq analysis of kidneys comparing gene expression profiles in black bears entering (late fall) and emerging (early spring) from hibernation identified 169 protein-coding genes that were differentially expressed. Of these, 101 genes were downregulated and 68 genes were upregulated after hibernation. Fold changes ranged from 1.8-fold downregulation (RTN4RL2) to 2.4-fold upregulation (CISH). Most notable was the upregulation of cytokine suppression genes (SOCS2, CISH, and SERPINC1) and the lack of increased expression of cytokines and genes involved in inflammation. The identification of these differences in gene expression in the black bear kidney may provide new insights in the prevention and treatment of CKD. © The Author(s) 2018. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.

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April 21, 2020

Potential for Adaptation to Climate Change Through Genomic Breeding in Sesame

Sesame is an important oilseed crop with high oil content and oil quality. Abundant unsaturated fatty acids, proteins, and antioxidants in sesame seeds attract the worldwide consumption of sesame products. Sesame is highly tolerant of drought and poor soil condition, even though it is readily affected by diseases and waterlogging stress, thereby leading to reduced seed yield and quality. For sesame, increasing the high and stable yield is requisite and urgent. Meanwhile, it is necessary to increase the mechanization level of its harvest for the world’s sesame production. Sesame, S. indicum, is the sole cultivated species in Sesamum genus. The relatively low genetic diversity limits sesame breeding for new and substantial improved varieties. In this section, we present a review of the key agronomic traits and the breeding methods currently used in the species. We also pinpoint the achievement of the Sesame Genome Project (SGP) and the potential for the genomics-assisted breeding in sesame.

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April 21, 2020

Secretion of an Argonaute protein by a parasitic nematode and the evolution of its siRNA guides.

Extracellular RNA has been proposed to mediate communication between cells and organisms however relatively little is understood regarding how specific sequences are selected for export. Here, we describe a specific Argonaute protein (exWAGO) that is secreted in extracellular vesicles (EVs) released by the gastrointestinal nematode Heligmosomoides bakeri, at multiple copies per EV. Phylogenetic and gene expression analyses demonstrate exWAGO orthologues are highly conserved and abundantly expressed in related parasites but highly diverged in free-living genus Caenorhabditis. We show that the most abundant small RNAs released from the nematode parasite are not microRNAs as previously thought, but rather secondary small interfering RNAs (siRNAs) that are produced by RNA-dependent RNA Polymerases. The siRNAs that are released in EVs have distinct evolutionary properties compared to those resident in free-living or parasitic nematodes. Immunoprecipitation of exWAGO demonstrates that it specifically associates with siRNAs from transposons and newly evolved repetitive elements that are packaged in EVs and released into the host environment. Together this work demonstrates molecular and evolutionary selectivity in the small RNA sequences that are released in EVs into the host environment and identifies a novel Argonaute protein as the mediator of this. © The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.

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April 21, 2020

A 12-kb structural variation in progressive myoclonic epilepsy was newly identified by long-read whole-genome sequencing.

We report a family with progressive myoclonic epilepsy who underwent whole-exome sequencing but was negative for pathogenic variants. Similar clinical courses of a devastating neurodegenerative phenotype of two affected siblings were highly suggestive of a genetic etiology, which indicates that the survey of genetic variation by whole-exome sequencing was not comprehensive. To investigate the presence of a variant that remained unrecognized by standard genetic testing, PacBio long-read sequencing was performed. Structural variant (SV) detection using low-coverage (6×) whole-genome sequencing called 17,165 SVs (7,216 deletions and 9,949 insertions). Our SV selection narrowed down potential candidates to only five SVs (two deletions and three insertions) on the genes tagged with autosomal recessive phenotypes. Among them, a 12.4-kb deletion involving the CLN6 gene was the top candidate because its homozygous abnormalities cause neuronal ceroid lipofuscinosis. This deletion included the initiation codon and was found in a GC-rich region containing multiple repetitive elements. These results indicate the presence of a causal variant in a difficult-to-sequence region and suggest that such variants that remain enigmatic after the application of current whole-exome sequencing technology could be uncovered by unbiased application of long-read whole-genome sequencing.

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April 21, 2020

Fast and accurate genomic analyses using genome graphs.

The human reference genome serves as the foundation for genomics by providing a scaffold for alignment of sequencing reads, but currently only reflects a single consensus haplotype, thus impairing analysis accuracy. Here we present a graph reference genome implementation that enables read alignment across 2,800 diploid genomes encompassing 12.6 million SNPs and 4.0 million insertions and deletions (indels). The pipeline processes one whole-genome sequencing sample in 6.5?h using a system with 36?CPU cores. We show that using a graph genome reference improves read mapping sensitivity and produces a 0.5% increase in variant calling recall, with unaffected specificity. Structural variations incorporated into a graph genome can be genotyped accurately under a unified framework. Finally, we show that iterative augmentation of graph genomes yields incremental gains in variant calling accuracy. Our implementation is an important advance toward fulfilling the promise of graph genomes to radically enhance the scalability and accuracy of genomic analyses.

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April 21, 2020

The CF Canada-Sick Kids Program in individual CF therapy: A resource for the advancement of personalized medicine in CF.

Therapies targeting certain CFTR mutants have been approved, yet variations in clinical response highlight the need for in-vitro and genetic tools that predict patient-specific clinical outcomes. Toward this goal, the CF Canada-Sick Kids Program in Individual CF Therapy (CFIT) is generating a “first of its kind”, comprehensive resource containing patient-specific cell cultures and data from 100 CF individuals that will enable modeling of therapeutic responses.The CFIT program is generating: 1) nasal cells from drug naïve patients suitable for culture and the study of drug responses in vitro, 2) matched gene expression data obtained by sequencing the RNA from the primary nasal tissue, 3) whole genome sequencing of blood derived DNA from each of the 100 participants, 4) induced pluripotent stem cells (iPSCs) generated from each participant’s blood sample, 5) CRISPR-edited isogenic control iPSC lines and 6) prospective clinical data from patients treated with CF modulators.To date, we have recruited 57 of 100 individuals to CFIT, most of whom are homozygous for F508del (to assess in-vitro: in-vivo correlations with respect to ORKAMBI response) or heterozygous for F508del and a minimal function mutation. In addition, several donors are homozygous for rare nonsense and missense mutations. Nasal epithelial cell cultures and matched iPSC lines are available for many of these donors.This accessible resource will enable development of tools that predict individual outcomes to current and emerging modulators targeting F508del-CFTR and facilitate therapy discovery for rare CF causing mutations.Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

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April 21, 2020

rMETL: sensitive mobile element insertion detection with long read realignment.

Mobile element insertion (MEI) is a major category of structure variations (SVs). The rapid development of long read sequencing technologies provides the opportunity to detect MEIs sensitively. However, the signals of MEI implied by noisy long reads are highly complex due to the repetitiveness of mobile elements as well as the high sequencing error rates. Herein, we propose the Realignment-based Mobile Element insertion detection Tool for Long read (rMETL). Benchmarking results of simulated and real datasets demonstrate that rMETL enables to handle the complex signals to discover MEIs sensitively. It is suited to produce high-quality MEI callsets in many genomics studies.rMETL is available from https://github.com/hitbc/rMETL.Supplementary data are available at Bioinformatics online. © The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

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April 21, 2020

Genome Sequence of Jaltomata Addresses Rapid Reproductive Trait Evolution and Enhances Comparative Genomics in the Hyper-Diverse Solanaceae.

Within the economically important plant family Solanaceae, Jaltomata is a rapidly evolving genus that has extensive diversity in flower size and shape, as well as fruit and nectar color, among its ~80 species. Here, we report the whole-genome sequencing, assembly, and annotation, of one representative species (Jaltomata sinuosa) from this genus. Combining PacBio long reads (25×) and Illumina short reads (148×) achieved an assembly of ~1.45?Gb, spanning ~96% of the estimated genome. Ninety-six percent of curated single-copy orthologs in plants were detected in the assembly, supporting a high level of completeness of the genome. Similar to other Solanaceous species, repetitive elements made up a large fraction (~80%) of the genome, with the most recently active element, Gypsy, expanding across the genome in the last 1-2 Myr. Computational gene prediction, in conjunction with a merged transcriptome data set from 11 tissues, identified 34,725 protein-coding genes. Comparative phylogenetic analyses with six other sequenced Solanaceae species determined that Jaltomata is most likely sister to Solanum, although a large fraction of gene trees supported a conflicting bipartition consistent with substantial introgression between Jaltomata and Capsicum after these species split. We also identified gene family dynamics specific to Jaltomata, including expansion of gene families potentially involved in novel reproductive trait development, and loss of gene families that accompanied the loss of self-incompatibility. This high-quality genome will facilitate studies of phenotypic diversification in this rapidly radiating group and provide a new point of comparison for broader analyses of genomic evolution across the Solanaceae.

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April 21, 2020

Heterochromatin-enriched assemblies reveal the sequence and organization of the Drosophila melanogaster Y chromosome.

Heterochromatic regions of the genome are repeat-rich and poor in protein coding genes, and are therefore underrepresented in even the best genome assemblies. One of the most difficult regions of the genome to assemble are sex-limited chromosomes. The Drosophila melanogaster Y chromosome is entirely heterochromatic, yet has wide-ranging effects on male fertility, fitness, and genome-wide gene expression. The genetic basis of this phenotypic variation is difficult to study, in part because we do not know the detailed organization of the Y chromosome. To study Y chromosome organization in D. melanogaster, we develop an assembly strategy involving the in silico enrichment of heterochromatic long single-molecule reads and use these reads to create targeted de novo assemblies of heterochromatic sequences. We assigned contigs to the Y chromosome using Illumina reads to identify male-specific sequences. Our pipeline extends the D. melanogaster reference genome by 11.9 Mb, closes 43.8% of the gaps, and improves overall contiguity. The addition of 10.6 MB of Y-linked sequence permitted us to study the organization of repeats and genes along the Y chromosome. We detected a high rate of duplication to the pericentric regions of the Y chromosome from other regions in the genome. Most of these duplicated genes exist in multiple copies. We detail the evolutionary history of one sex-linked gene family, crystal-Stellate While the Y chromosome does not undergo crossing over, we observed high gene conversion rates within and between members of the crystal-Stellate gene family, Su(Ste), and PCKR, compared to genome-wide estimates. Our results suggest that gene conversion and gene duplication play an important role in the evolution of Y-linked genes. Copyright © 2019 Chang and Larracuente.

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April 21, 2020

Effective approaches to study the plant-root knot nematode interaction.

Plant-parasitic nematodes cause major agricultural losses worldwide. Examining the molecular mechanisms underlying plant-nematode interactions and how plants respond to different invading pathogens is attracting major attention to reduce the expanding gap between agricultural production and the needs of the growing world population. This review summarizes the most recent developments in plant-nematode interactions and the diverse approaches used to improve plant resistance against root knot nematode (RKN). We will emphasize the recent rapid advances in genome sequencing technologies, small interfering RNA techniques (RNAi) and targeted genome editing which are contributing to the significant progress in understanding the plant-nematode interaction mechanisms. Also, molecular approaches to improve plant resistance against nematodes are considered.Copyright © 2019 Elsevier Masson SAS. All rights reserved.

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April 21, 2020

Short communication: Identification of the pseudoautosomal region in the Hereford bovine reference genome assembly ARS-UCD1.2.

In cattle, the X chromosome accounts for approximately 3 and 6% of the genome in bulls and cows, respectively. In spite of the large size of this chromosome, very few studies report analysis of the X chromosome in genome-wide association studies and genomic selection. This lack of genetic interrogation is likely due to the complexities of undertaking these studies given the hemizygous state of some, but not all, of the X chromosome in males. The first step in facilitating analysis of this gene-rich chromosome is to accurately identify coordinates for the pseudoautosomal boundary (PAB) to split the chromosome into a region that may be treated as autosomal sequence (pseudoautosomal region) and a region that requires more complex statistical models. With the recent release of ARS-UCD1.2, a more complete and accurate assembly of the cattle genome than was previously available, it is timely to fine map the PAB for the first time. Here we report the use of SNP chip genotypes, short-read sequences, and long-read sequences to fine map the PAB (X chromosome:133,300,518) and simultaneously determine the neighboring regions of reduced homology and true pseudoautosomal region. These results greatly facilitate the inclusion of the X chromosome in genome-wide association studies, genomic selection, and other genetic analysis undertaken on this reference genome.The Authors. Published by FASS Inc. and Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

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April 21, 2020

Breeding Brassica juncea and B. rapa for Sustainable Oilseed Production in the Changing Climate: Progress and Prospects

The uncertainties of climatic variability and global warming are leading to rising concerns towards ensuring global food security of an expanding population. Unfavorable climatic conditions, like extremes of temperature, drought, flood, and salinity, in addition to the elevated greenhouse gases adversely affect the physiology, and accordingly the quantitative and qualitative characteristics of plants. Mustard (Brassica juncea) and rape (Brassica rapa), the two important oilseed crops of the Indian subcontinent, are also cultivated in Eastern Europe, Russia, China, and Canada. These oilseed crops are affected by various biotic and abiotic stress during different growth and developmental stages, that severely influences agricultural productivity. Extensive breeding efforts toward the development of Brassica cultivars that can resist these climatic variabilities are under various stages of progress. The Brassica germplasm and the wild relatives of B. juncea and B. rapa, which constitute important genetic stocks, are also being utilized in these breeding programs. An integrated approach is required that will study plant-insect pest and disease-climate interactions for conceiving future strategies to develop disease-, insect-resistant, and climate-resilient plant varieties. Developing mustard varieties, efficient in the utilization of soil nutrients, are also required for improving productivity in impoverished soils and for  better uptake/utilization of nutrients in soils rich in resources. Future research in oilseed mustard and rape should, therefore, involve examining the influence of climate-smart traits on yield/production in targeted environments, so that climate-resilient cultivars adapted to climate change conditions could be developed. This chapter summarizes the advances in breeding of climate-smart traits such as,  tolerance to drought, heat, salinity, flooding and frost, and efficient nutrient utilization, in oilseed mustard and rape, that could assist in the genomic designing for climate-smart crops.

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April 21, 2020

Smashing Barriers in Biolistic Plant Transformation.

A foundation of modern biotechnology is the ability to stably introduce foreign DNA into an organism. The two most widely used methods, Agrobacterium-mediated transformation and biolistics, are both steeped in a rich history of creative exploration into the molecular unknown. Agrobacterium research accelerated in the early 1970s, particularly with the discovery of the large Ti (tumor-inducing) plasmid of Agrobacterium that contained a region of transfer DNA (T-DNA). Culturing plant calli in autoclaved jelly jars, and long before the advent of PCR, Southern blots were first used to show that T-DNA fragments could stably integrate into the nuclear genome (Chilton et al., 1980; Chilton, 2001). On the other hand, the first manufactured biolistic “gene gun” was an actual gun; it shot a blank .22 caliber cartridge loaded with DNA-coated tungsten shards to integrate foreign DNA into the nuclear genome. While it has long been known that biolistic transformation violently integrates DNA in a largely random, unpredictable and imprecise way, the cellular mechanisms of damage repair and successful integration remain a complicated issue to disentangle.

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