De Novo Hepatic Transcriptome Assembly and Systems Level Analysis of Three Species of Dietary Fish, Sardinops sagax, Scomber japonicus, and Pleuronichthys verticalis

The monitoring of marine species as sentinels for ecosystem health has long been a valuable tool worldwide, providing insight into how both anthropogenic pollution and naturally occurring phenomena (i.e., harmful algal blooms) may lead to human and animal dietary concerns. The marine environments contain many contaminants of anthropogenic origin that have sufficient similarities to steroid and thyroid hormones, to potentially disrupt normal endocrine physiology in humans, fish, and other animals. An appropriate understanding of the effects of these endocrine disrupting chemicals (EDCs) on forage fish (e.g., sardine, anchovy, mackerel) can lead to significant insight into how these contaminants may affect local ecosystems in addition to their potential impacts on human health. With advancements in molecular tools (e.g., high-throughput sequencing, HTS), a genomics approach offers a robust toolkit to discover putative genetic biomarkers in fish exposed to these chemicals. However, the lack of available sequence information for non-model species has limited the development of these genomic toolkits. Using HTS and de novo assembly technology, the present study aimed to establish, for the first time for Sardinops sagax (Pacific sardine), Scomber japonicas (Pacific chub mackerel) and Pleuronichthys verticalis (hornyhead turbot), a de novo global transcriptome database of the liver, the primary organ involved in detoxification. The assembled transcriptomes provide a foundation for further downstream validation, comparative genomic analysis and biomarker development for future applications in ecotoxicogenomic studies, as well as environmental evaluation (e.g., climate change) and public health safety (e.g., dietary screening).

Download Full-text

Assessing Genome-Wide Diversity in European Hantaviruses through Sequence Capture from Natural Host Samples

Viruses ◽

10.3390/v12070749 ◽

2020 ◽

Vol 12 (7) ◽

pp. 749 ◽

Cited By ~ 3

Author(s):

Melanie Hiltbrunner ◽

Gerald Heckel

Keyword(s):

High Throughput Sequencing ◽

De Novo ◽

Phylogenetic Analyses ◽

Common Vole ◽

Natural Host ◽

Sequence Information ◽

Coding Region ◽

Sequence Capture ◽

Genome Wide ◽

The Impact

Research on the ecology and evolution of viruses is often hampered by the limitation of sequence information to short parts of the genomes or single genomes derived from cultures. In this study, we use hybrid sequence capture enrichment in combination with high-throughput sequencing to provide efficient access to full genomes of European hantaviruses from rodent samples obtained in the field. We applied this methodology to Tula (TULV) and Puumala (PUUV) orthohantaviruses for which analyses from natural host samples are typically restricted to partial sequences of their tri-segmented RNA genome. We assembled a total of ten novel hantavirus genomes de novo with very high coverage (on average >99%) and sequencing depth (average >247×). A comparison with partial Sanger sequences indicated an accuracy of >99.9% for the assemblies. An analysis of two common vole (Microtus arvalis) samples infected with two TULV strains each allowed for the de novo assembly of all four TULV genomes. Combining the novel sequences with all available TULV and PUUV genomes revealed very similar patterns of sequence diversity along the genomes, except for remarkably higher diversity in the non-coding region of the S-segment in PUUV. The genomic distribution of polymorphisms in the coding sequence was similar between the species, but differed between the segments with the highest sequence divergence of 0.274 for the M-segment, 0.265 for the S-segment, and 0.248 for the L-segment (overall 0.258). Phylogenetic analyses showed the clustering of genome sequences consistent with their geographic distribution within each species. Genome-wide data yielded extremely high node support values, despite the impact of strong mutational saturation that is expected for hantavirus sequences obtained over large spatial distances. We conclude that genome sequencing based on capture enrichment protocols provides an efficient means for ecological and evolutionary investigations of hantaviruses at an unprecedented completeness and depth.

Download Full-text

De novo assembly and inferred functional annotation of the transcriptome of Heterosigma akashiwo

10.22541/au.164112034.43396317/v1 ◽

2022 ◽

Author(s):

Masanao Sato ◽

Masahide Seki ◽

Yutaka Suzuki ◽

Shoko Ueki

Keyword(s):

Functional Annotation ◽

De Novo ◽

Molecular Level ◽

Transcriptome Assembly ◽

Practical Interest ◽

Heterosigma Akashiwo ◽

Sequence Information ◽

Biological Characterization ◽

Nucleotide Database

Heterosigma akashiwo is a eukaryotic, cosmopolitan, and unicellular alga (class: Raphidophyceae), and produces fish-killing blooms. There is a substantial scientific and practical interest in its ecophysiological characteristics that determine bloom dynamics and its adaptation to broad climate zones. A well-annotated genomic/genetic sequence information enables researchers to characterize organisms using modern molecular technology. The Chloroplast and the mitochondrial genome sequences and transcriptome sequence assembly (TSA) datasets with limited sizes for H. akashiwo are available in NCBI nucleotide database on December 2021: there is no doubt that more genetic information of the species will greatly enhance the progress of biological characterization of the species. Here, we conducted H. akashiwo RNA sequencing, a de novo transcriptome assembly (NCBI TSA ICRV01) of a large number of high-quality short-read sequences, and the functional annotation of predicted genes. Based on our transcriptome, we confirmed that the organism possesses genes that were predicted to function in phagocytosis, supporting the earlier observations of H. akashiwo bacterivory. Along with its capability for photosynthesis, the mixotrophy of H. akashiwo may partially explain its high adaptability to various environmental conditions. Our study here will provide an important toehold to decipher H. akashiwo ecophysiology at a molecular level.

Download Full-text

Characterization of the transcriptome and EST-SSR development in Boea clarkeana, a desiccation-tolerant plant endemic to China

10.7287/peerj.preprints.2603v1 ◽

2016 ◽

Author(s):

Ying Wang ◽

Kun Liu ◽

De Bi ◽

Biao Shou Zhou ◽

Wen Jian Shao

Keyword(s):

De Novo ◽

Gene Annotation ◽

Sequence Similarity ◽

Molecular Study ◽

Sequence Information ◽

Sequencing Data ◽

Protein Database ◽

Illumina Hiseq ◽

Significant Similarity ◽

Assembly Technology

Background. Resurrection plants constitute a unique cadre within angiosperms. Boea clarkeana Hemsl. (Boea, Gesneriaceae) is a desiccation-tolerant dicotyledonous herb that is endemic to China. Although research on angiosperms with DT could be instructive for crops, genomic resources for B. clarkeana remain scarce. In addition, transcriptome sequencing could be an effective way to study desiccation-tolerant plants. Methods. In the present study, we used the platform Illumina HiSeqTM 2000 and de novo assembly technology to obtain leaf transcriptomes of B. clarkeana and conducted a BLASTX alignment of the sequencing data and protein databases for sequence classification and annotation. Then, based on the sequence information obtained, we developed EST-SSR markers by means of EST-SSR mining, primer design and polymorphism identification. Results. A total of 91,449 unigenes were generated from the leaf cDNA library of B. clarkeana in this study. Based on a sequence similarity search with a known protein database, 72,087 unigenes were annotated. Among the annotated unigenes, a total of 71,170 unigenes showed significant similarity to known proteins of 463 popular model species in the Nr database, and 59,962 unigenes and 32,336 unigenes were assigned to GO classifications and COG, respectively. In addition, 44,924 unigenes were mapped in 128 KEGG pathways. Furthermore, a total of 7,610 unigenes with 8,563 microsatellites were found. Seventy-four primer pairs were selected from 436 primer pairs designed for polymorphism validation. SSRs with higher polymorphism rates were concentrated on dinucleotides, pentanucleotides and hexanucleotides. Finally, 17 pairs with highly polymorphic and stable loci were selected for polymorphism screening. There were a total of 65 alleles, with 2–6 alleles at each locus. Mainly due to the unique biological characteristics of plants, the HE, HO and PIC per locus were very low, ranging from 0 to 0.196, 0.082 to 0.14 and 0 to 0.155, respectively. Discussion. A substantial fraction transcriptome sequences of B. clarkeana were generated in this study, which is the first molecular-level analysis of this plant. These sequences are valuable resources for gene annotation and discovery and molecular marker development. These sequences could also provide a valuable basis for the future molecular study of B. clarkeana.

Download Full-text

Proteotranscriptomics assisted gene annotation and spatial proteomics of Bombyx mori BmN4 cell line

10.21203/rs.3.rs-23159/v2 ◽

2020 ◽

Author(s):

Michal Levin ◽

Marion Scheibe ◽

Falk Butter

Keyword(s):

Mass Spectrometry ◽

Bombyx Mori ◽

Cell Line ◽

De Novo ◽

High Resolution Mass Spectrometry ◽

Gene Annotation ◽

Transcriptome Assembly ◽

Model Organisms ◽

Sequence Information ◽

A Genome

Abstract BackgroundThe process of identifying all coding regions in a genome is crucial for any study at the level of molecular biology, ranging from single-gene cloning to genome-wide measurements using RNA-Seq or mass spectrometry. While satisfactory annotation has been made feasible for well-studied model organisms through great efforts of big consortia, for most systems this kind of data is either absent or not adequately precise. ResultsCombining in-depth transcriptome sequencing and high resolution mass spectrometry, we here use proteotranscriptomics to improve gene annotation of protein-coding genes in the Bombyx mori cell line BmN4 which is an increasingly used tool for the analysis of piRNA biogenesis and function. Using this approach we provide the exact coding sequence and evidence for more than 6,200 genes on the protein level. Furthermore using spatial proteomics, we establish the subcellular localization of thousands of these proteins. We show that our approach outperforms current Bombyx mori annotation attempts in terms of accuracy and coverage. ConclusionsWe show that proteotranscriptomics is an efficient, cost-effective and accurate approach to improve previous annotations or generate new gene models. As this technique is based on de-novo transcriptome assembly, it provides the possibility to study any species also in the absence of genome sequence information for which proteogenomics would be impossible.

Download Full-text

Genome skimming reveals novel plastid markers for the molecular identification of illegally logged African timber species

PLoS ONE ◽

10.1371/journal.pone.0251655 ◽

2021 ◽

Vol 16 (6) ◽

pp. e0251655

Author(s):

Maurizio Mascarello ◽

Mario Amalfi ◽

Pieter Asselman ◽

Erik Smets ◽

Olivier J. Hardy ◽

...

Keyword(s):

Related Species ◽

High Throughput Sequencing ◽

De Novo ◽

Microscopic Analysis ◽

Comparative Genomic ◽

Timber Species ◽

Illegal Logging ◽

Closely Related Species ◽

Genome Skimming ◽

The Tropics

Tropical forests represent vast carbon stocks and continue to be key carbon sinks and buffer climate changes. The international policy constructed several mechanisms aiming at conservation and sustainable use of these forests. Illegal logging is an important threat of forests, especially in the tropics. Several laws and regulations have been set up to combat illegal timber trade. Despite significant enforcement efforts of these regulations, illegal logging continues to be a serious problem and impacts for the functioning of the forest ecosystem and global biodiversity in the tropics. Microscopic analysis of wood samples and the use of conventional plant DNA barcodes often do not allow to distinguish closely-related species. The use of novel molecular technologies could make an important contribution for the identification of tree species. In this study, we used high-throughput sequencing technologies and bioinformatics tools to obtain the complete de-novo chloroplast genome of 62 commercial African timber species using the genome skimming method. Then, we performed a comparative genomic analysis that revealed new candidate genetic regions for the discrimination of closely-related species. We concluded that genome skimming is a promising method for the development of plant genetic markers to combat illegal logging activities supporting CITES, FLEGT and the EU Timber Regulation.

Download Full-text

De novo assembly and inferred functional annotation of the transcriptome of Heterosigma akashiwo

10.22541/au.164112034.43396317/v2 ◽

2022 ◽

Author(s):

Masanao Sato ◽

Masahide Seki ◽

Yutaka Suzuki ◽

Shoko Ueki

Keyword(s):

Functional Annotation ◽

De Novo ◽

Molecular Level ◽

Transcriptome Assembly ◽

Practical Interest ◽

Heterosigma Akashiwo ◽

Sequence Information ◽

Biological Characterization ◽

Nucleotide Database

Download Full-text

A de novo 2.78-Mb duplication on chromosome 21q22.11 implicates candidate genes in the partial trisomy 21 phenotype

npj Genomic Medicine ◽

10.1038/npjgenmed.2016.3 ◽

2016 ◽

Vol 1 (1) ◽

Cited By ~ 5

Author(s):

James D Weisfeld-Adams ◽

Amanda K Tkachuk ◽

Kenneth N Maclean ◽

Naomi L Meeks ◽

Stuart A Scott

Keyword(s):

Candidate Genes ◽

Clinical Features ◽

Critical Region ◽

Trisomy 21 ◽

High Throughput Sequencing ◽

De Novo ◽

Partial Trisomy ◽

Chromosome 21 ◽

Comparative Genomic Hybridisation ◽

Comparative Genomic

Abstract Down syndrome (DS) is the most common genetic cause of intellectual disability (ID) and in the majority of cases is the result of complete trisomy 21. The hypothesis that the characteristic DS clinical features are due to a single DS critical region (DSCR) at distal chromosome 21q has been refuted by recently reported segmental trisomy 21 cases characterised by microarray-based comparative genomic hybridisation (aCGH). These rare cases have implicated multiple regions on chromosome 21 in the aetiology of distinct features of DS; however, the map of chromosome 21 copy-number aberrations and their associated phenotypes remains incompletely defined. We report a child with ID who was deemed very high risk for DS on antenatal screening (1 in 13) and has partial, but distinct, dysmorphologic features of DS without congenital heart disease (CHD). Oligonucleotide aCGH testing of the proband detected a previously unreported de novo 2.78-Mb duplication on chromosome 21q22.11 that includes 16 genes; however, this aberration does not harbour any of the historical DSCR genes (APP, DSCR1, DYRK1A and DSCAM). This informative case implicates previously under-recognised candidate genes (SOD1, SYNJ1 and ITSN1) in the pathogenesis of specific DS clinical features and supports a critical region for CHD located more distal on chromosome 21q. In addition, this unique case illustrates how the increasing resolution of microarray and high-throughput sequencing technologies can continue to reveal new biology and enhance understanding of widely studied genetic diseases that were originally described over 50 years ago.

Download Full-text

High-throughput sequencing and de novo transcriptome assembly of Swertia japonica to identify genes involved in the biosynthesis of therapeutic metabolites

Plant Cell Reports ◽

10.1007/s00299-016-2021-z ◽

2016 ◽

Vol 35 (10) ◽

pp. 2091-2111 ◽

Cited By ~ 20

Author(s):

Amit Rai ◽

Michimi Nakamura ◽

Hiroki Takahashi ◽

Hideyuki Suzuki ◽

Kazuki Saito ◽

...

Keyword(s):

High Throughput ◽

High Throughput Sequencing ◽

De Novo ◽

Transcriptome Assembly ◽

De Novo Transcriptome Assembly ◽

De Novo Transcriptome ◽

Swertia Japonica

Download Full-text

De novo transcriptome assembly of the Italian white truffle (Tuber magnatum Pico)

10.1101/461483 ◽

2018 ◽

Cited By ~ 1

Author(s):

Federico Vita ◽

Amedeo Alpi ◽

Edoardo Bertolini

Keyword(s):

Molecular Mechanisms ◽

De Novo ◽

Transcriptome Assembly ◽

Rna Seq ◽

Genetic Studies ◽

Protein Coding ◽

Important Species ◽

White Truffle ◽

Tuber Magnatum ◽

Insight Into

AbstractThe Italian white truffle (Tuber magnatum Pico) is a gastronomic delicacy that dominates the worldwide truffle market. Despite its importance, the genomic resources currently available for this species are still limited. Here we present the first de novo transcriptome assembly of T. magnatum. Illumina RNA-seq data were assembled using a single-k-mer approach into 22,932 transcripts with N50 of 1,524 bp. Our approach allowed to predict and annotate 12,367 putative protein coding sequences, reunited in 6,723 loci. In addition, we identified 2,581 gene-based SSR markers. This work provides the first publicly available reference transcriptome for genomics and genetic studies providing insight into the molecular mechanisms underlying the biology of this important species.

Download Full-text