scholarly journals De NovoAssembly of the Pea (Pisum sativumL.) Nodule Transcriptome

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Vladimir A. Zhukov ◽  
Alexander I. Zhernakov ◽  
Olga A. Kulaeva ◽  
Nikita I. Ershov ◽  
Alexey Y. Borisov ◽  
...  
Keyword(s):  
Transcriptome Sequencing ◽  
De Novo ◽  
Transcriptome Assembly ◽  
Specific Gene ◽  
Its Sequencing ◽  
Sequencing Data ◽  
Root Tips ◽  
Sequencing Project ◽  
Illumina Gaiix ◽  
Project Data

The large size and complexity of the garden pea (Pisum sativumL.) genome hamper its sequencing and the discovery of pea gene resources. Although transcriptome sequencing provides extensive information about expressed genes, some tissue-specific transcripts can only be identified from particular organs under appropriate conditions. In this study, we performed RNA sequencing of polyadenylated transcripts from young pea nodules and root tips on an Illumina GAIIx system, followed byde novotranscriptome assembly using the Trinity program. We obtained more than 58,000 and 37,000 contigs from “Nodules” and “Root Tips” assemblies, respectively. The quality of the assemblies was assessed by comparison with pea expressed sequence tags and transcriptome sequencing project data available from NCBI website. The “Nodules” assembly was compared with the “Root Tips” assembly and with pea transcriptome sequencing data from projects indicating tissue specificity. As a result, approximately 13,000 nodule-specific contigs were found and annotated by alignment to known plant protein-coding sequences and by Gene Ontology searching. Of these, 581 sequences were found to possess full CDSs and could thus be considered as novel nodule-specific transcripts of pea. The information about pea nodule-specific gene sequences can be applied for gene-based markers creation, polymorphism studies, and real-time PCR.

scholarly journals RNA-Bloom provides lightweight reference-free transcriptome assembly for single cells

2019 ◽  
Author(s):  
Ka Ming Nip ◽  
Readman Chiu ◽  
Chen Yang ◽  
Justin Chu ◽  
Hamid Mohamadi ◽  
...  
Keyword(s):  
De Novo ◽  
Transcriptome Assembly ◽  
Single Cells ◽  
Specific Gene ◽  
Rna Seq ◽  
Sequencing Data ◽  
Assembly Algorithm ◽  
The Rich ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome

We present RNA-Bloom, a de novo RNA-seq assembly algorithm that leverages the rich information content in single-cell transcriptome sequencing (scRNA-seq) data to reconstruct cell-specific isoforms. We benchmark RNA-Bloom’s performance against leading bulk RNA-seq assembly approaches, and illustrate its utility in detecting cell-specific gene fusion events using sequencing data from HiSeq-4000 and BGISEQ-500 platforms. We expect RNA-Bloom to boost the utility of scRNA-seq data, expanding what is informatically accessible now.

Computational Approaches for Transcriptome Assembly Based on Sequencing Technologies

2020 ◽  
Vol 15 (1) ◽  
pp. 2-16
Author(s):  
Yuwen Luo ◽  
Xingyu Liao ◽  
Fang-Xiang Wu ◽  
Jianxin Wang
Keyword(s):  
De Novo ◽  
Transcriptome Assembly ◽  
Critical Role ◽  
High Sensitivity ◽  
Biological Properties ◽  
Sequencing Data ◽  
Sequencing Technologies ◽  
Long Reads ◽  
Massive Sequencing ◽  
Generation Sequencing

Transcriptome assembly plays a critical role in studying biological properties and examining the expression levels of genomes in specific cells. It is also the basis of many downstream analyses. With the increase of speed and the decrease in cost, massive sequencing data continues to accumulate. A large number of assembly strategies based on different computational methods and experiments have been developed. How to efficiently perform transcriptome assembly with high sensitivity and accuracy becomes a key issue. In this work, the issues with transcriptome assembly are explored based on different sequencing technologies. Specifically, transcriptome assemblies with next-generation sequencing reads are divided into reference-based assemblies and de novo assemblies. The examples of different species are used to illustrate that long reads produced by the third-generation sequencing technologies can cover fulllength transcripts without assemblies. In addition, different transcriptome assemblies using the Hybrid-seq methods and other tools are also summarized. Finally, we discuss the future directions of transcriptome assemblies.

Identification of tissue-specific gene clusters and orthologues of nodulation-related genes in Vigna angularis

2014 ◽  
Vol 12 (S1) ◽  
pp. S21-S26 ◽  
Author(s):  
Yang Jae Kang ◽  
Jayern Lee ◽  
Yong Hwan Kim ◽  
Suk-Ha Lee
Keyword(s):  
De Novo ◽  
Transcriptome Assembly ◽  
Lotus Japonicus ◽  
Gene Clusters ◽  
Specific Gene ◽  
Vigna Angularis ◽  
Model Legume ◽  
Tissue Specific ◽  
Autoregulation Of Nodulation ◽  
Species Specific

Nitrogen fixation in legumes is an important agricultural trait that results from symbiosis between the root and rhizobia. To understand the molecular basis of nodulation, recent research has been focused on the identification of nodulation-related genes by functional analysis using two major model legumes, Medicago truncatula and Lotus japonicus. Thus far, three important processes have been discovered, namely Nod factor (NF) perception, NF signalling and autoregulation of nodulation. Nevertheless, application of the results of these studies is limited for non-model legume crops because a reference genome is unavailable. However, because the cost of whole-transcriptome analysis has dropped dramatically due to the Next generation sequencer (NGS) technology, minor crops for which reference sequences are yet to be constructed can still be studied at the genome level. In this study, we sequenced the leaf and root transcriptomes of Vigna angularis (accession IT213134) and de novo assembled. Our results demonstrate the feasibility of using the transcriptome assembly to effectively identify tissue-specific peptide clusters related to tissue-specific functions and species-specific nodulation-related genes.

scholarly journals Raw transcriptomics data to gene specific SSRs: a validated free bioinformatics workflow for biologists

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
D. N. U. Naranpanawa ◽  
C. H. W. M. R. B. Chandrasekara ◽  
P. C. G. Bandaranayake ◽  
A. U. Bandaranayake
Keyword(s):  
De Novo ◽  
Sequence Data ◽  
Transcriptome Assembly ◽  
Low Cost ◽  
Santalum Album ◽  
Sequencing Data ◽  
Illumina Hiseq ◽  
Tissue Samples ◽  
Downstream Analysis ◽  
Bioinformatics Workflow

Abstract Recent advances in next-generation sequencing technologies have paved the path for a considerable amount of sequencing data at a relatively low cost. This has revolutionized the genomics and transcriptomics studies. However, different challenges are now created in handling such data with available bioinformatics platforms both in assembly and downstream analysis performed in order to infer correct biological meaning. Though there are a handful of commercial software and tools for some of the procedures, cost of such tools has made them prohibitive for most research laboratories. While individual open-source or free software tools are available for most of the bioinformatics applications, those components usually operate standalone and are not combined for a user-friendly workflow. Therefore, beginners in bioinformatics might find analysis procedures starting from raw sequence data too complicated and time-consuming with the associated learning-curve. Here, we outline a procedure for de novo transcriptome assembly and Simple Sequence Repeats (SSR) primer design solely based on tools that are available online for free use. For validation of the developed workflow, we used Illumina HiSeq reads of different tissue samples of Santalum album (sandalwood), generated from a previous transcriptomics project. A portion of the designed primers were tested in the lab with relevant samples and all of them successfully amplified the targeted regions. The presented bioinformatics workflow can accurately assemble quality transcriptomes and develop gene specific SSRs. Beginner biologists and researchers in bioinformatics can easily utilize this workflow for research purposes.

scholarly journals A de novo Transcriptome Assembly of the European Flounder (Platichthys flesus): The Preselection of Transcripts Encoding Active Forms of Enzymes

2021 ◽  
Vol 8 ◽  
Author(s):  
Konrad Pomianowski ◽  
Artur Burzyński ◽  
Ewa Kulczykowska
Keyword(s):  
De Novo ◽  
Transcriptome Assembly ◽  
Head Kidney ◽  
Sequencing Data ◽  
Mrna Isoforms ◽  
Protein Database ◽  
Next Generation Sequencing Technology ◽  
Platichthys Flesus ◽  
European Flounder ◽  
High Level

The RNA sequencing data sets available for different fish species show a potentially high variety of forms of enzymes just in teleosts. This is primarily considered an effect of the first round of whole-genome duplication with mutations in duplicated genes (isozymes) and alternative splicing of mRNA (isoforms). However, the abundance of the mRNA transcript variants is not necessarily reflected in the abundance of active forms of proteins. We have investigated the transcriptional profiles of two enzymes, aralkylamine N-acetyltransferase (AANAT: EC 2.3.1.87) and N-acetylserotonin O-methyltransferase (ASMT: EC 2.1.1.4), in the eyeball, brain, intestines, spleen, heart, liver, head kidney, gonads, and skin of the European flounder (Platichthys flesus). High-throughput next-generation sequencing technology NovaSeq6000 was used to generate 500M sequencing reads. These were then assembled and filtered producing 75k reliable contigs. Gene ontology (GO) terms were assigned to the majority of annotated contigs/unigenes based on the results of PFAM, PANTHER, UniProt, and InterPro protein database searches. BUSCOs statistics for metazoa, vertebrata, and actinopterygii databases showed that the reported transcriptome represents a high level of completeness. In this article, we show how to preselect transcripts encoding the active enzymes (isozymes or isoforms), using AANAT and ASMT in the European flounder as the examples. The data can be used as a tool to design the experiments as well as a basis for discussion of diversity of enzyme forms and their physiological relevance in teleosts.

scholarly journals Transcriptome Sequencing of the Endangered Species Elongate Loach (Leptobotia elongata) From the Yangtze River: De novo Transcriptome Assembly, Annotation, Identification and Validation of EST-SSR Markers

2021 ◽  
Vol 8 ◽  
Author(s):  
Yunbang Zhang ◽  
Jian Gao ◽  
Yunhai Zhang ◽  
Yuanchao Zou ◽  
Xiaojuan Cao
Keyword(s):  
Endangered Species ◽  
Genetic Information ◽  
Transcriptome Sequencing ◽  
Yangtze River ◽  
Expressed Sequence Tag ◽  
De Novo ◽  
Transcriptome Assembly ◽  
Habitat Destruction ◽  
The Yangtze River ◽  
Simple Sequence

Elongate loach (Leptobotia elongata) is endemic to middle and upper reaches of the Yangtze River in China. Due to overfishing and habitat destruction, this loach has become an endangered species. So far, lack of reliable genetic information and molecular markers has hindered the conservation and utilization of elongate loach resources. Therefore, we here performed an Illumina sequencing and de novo transcriptome assembly in elongate loach, and then developed polymorphic simple sequence repeat markers (SSRs). After assembly, 51,185 unigenes were obtained, with an average length of 1,496 bp. A total of 23,901 expressed sequence tag-simple sequence repeats (EST-SSRs) were identified, distributing in 14,422 unigenes, with a distribution frequency of 28.18%. Out of 16,885 designed EST-SSR primers, 150 primers (3 or 4 base repetition-dominated) were synthesized for polymorphic EST-SSR development. Then, 52 polymorphic EST-SSRs were identified, with polymorphism information contents (PIC) ranging from 0.03 to 0.88 (average 0.54). In conclusion, this was the first report of transcriptome sequencing of elongate loach. Meanwhile, we developed a set of polymorphic EST-SSRs for the loach. This study will provide an important basis, namely genetic information and polymorphic SSRs, for further population genetics and breeding studies of this endangered and economic loach in China.

scholarly journals ClusTrast: a short read de novo transcript isoform assembler guided by clustered contigs

2022 ◽  
Author(s):  
Karl Johan Westrin ◽  
Warren W Kretzschmar ◽  
Olof Emanuelsson
Keyword(s):  
Alternative Splicing ◽  
Reference Genome ◽  
De Novo ◽  
Transcriptome Assembly ◽  
Sequencing Data ◽  
Short Read ◽  
Transcript Isoforms ◽  
Short Reads ◽  
Moderate Reduction ◽  
Eukaryotic Species

Motivation: Transcriptome assembly from RNA sequencing data in species without a reliable reference genome has to be performed de novo, but studies have shown that de novo methods often have inadequate reconstruction ability of transcript isoforms. This impedes the study of alternative splicing, in particular for lowly expressed isoforms. Result: We present the de novo transcript isoform assembler ClusTrast, which clusters a set of guiding contigs by similarity, aligns short reads to the guiding contigs, and assembles each clustered set of short reads individually. We tested ClusTrast on datasets from six eukaryotic species, and showed that ClusTrast reconstructed more expressed known isoforms than any of the other tested de novo assemblers, at a moderate reduction in precision. An appreciable fraction were reconstructed to at least 95% of their length. We suggest that ClusTrast will be useful for studying alternative splicing in the absence of a reference genome. Availability and implementation: The code and usage instructions are available at https://github.com/karljohanw/clustrast.

scholarly journals De novo transcriptome assembly from the gonads of a scleractinian coral, Euphyllia ancora: Molecular mechanisms underlying scleractinian gametogenesis

2020 ◽  
Author(s):  
Yi-Ling Chiu ◽  
Shinya Shikina ◽  
Yuki Yoshioka ◽  
Chuya Shinzato ◽  
Ching-Fong Chang
Keyword(s):  
Sperm Motility ◽  
Molecular Mechanisms ◽  
De Novo ◽  
Transcriptome Assembly ◽  
Expression Profiles ◽  
Molecular Characteristics ◽  
Specific Gene ◽  
Ecological Data ◽  
Developmental Phases ◽  
Upregulated Genes

Abstract Background: Sexual reproduction of scleractinians has captured the attention of researchers and the general public for decades. Although extensive ecological data has been acquired, underlying molecular and cellular mechanisms remain largely unknown. In this study, to better understand mechanisms underlying gametogenesis, we isolated ovaries and testes at different developmental phases from a gonochoric coral, Euphyllia ancora , and adopted a transcriptomic approach to reveal sex- and phase-specific gene expression profiles. In particular, we explored genes associated with oocyte development and maturation, spermiogenesis, sperm motility / capacitation, and fertilization.Results: 1.6 billion raw reads were obtained from 24 gonadal samples. De novo assembly of trimmed reads, and elimination of contigs derived from symbiotic dinoflagellates (Symbiodiniaceae) and other organisms yielded a reference E. ancora gonadal transcriptome of 35,802 contigs. Analysis of 4 developmental phases identified 2,023 genes that were differentially expressed during oogenesis and 678 during spermatogenesis. In premature/mature ovaries, 631 genes were specifically upregulated, with 538 in mature testes. Upregulated genes included those involved in gametogenesis, gamete maturation, sperm motility / capacitation, and fertilization in other metazoans, including humans. Meanwhile, a large number of genes without homology to sequences in the SWISS-PROT database were also observed among upregulated genes in premature / mature ovaries and mature testes.Conclusions: Our findings show that scleractinian gametogenesis shares many molecular characteristics with that of other metazoans, but it also possesses unique characteristics developed during cnidarian and/or scleractinian evolution. To the best of our knowledge, this study is the first to create a gonadal transcriptome assembly from any scleractinian. This study and associated datasets provide a foundation for future studies regarding gametogenesis and differences between male and female colonies from molecular and cellular perspectives. Furthermore, our transcriptome assembly will be a useful reference for future development of sex-specific and/or stage-specific germ cell markers that can be used in coral aquaculture and ecological studies.

scholarly journals De novo transcriptome assembly from the gonads of a scleractinian coral, Euphyllia ancora: molecular mechanisms underlying scleractinian gametogenesis

BMC Genomics ◽  
2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Yi-Ling Chiu ◽  
Shinya Shikina ◽  
Yuki Yoshioka ◽  
Chuya Shinzato ◽  
Ching-Fong Chang
Keyword(s):  
Sperm Motility ◽  
Molecular Mechanisms ◽  
De Novo ◽  
Transcriptome Assembly ◽  
Expression Profiles ◽  
Molecular Characteristics ◽  
Specific Gene ◽  
Ecological Data ◽  
Developmental Phases ◽  
Upregulated Genes

Abstract Background Sexual reproduction of scleractinians has captured the attention of researchers and the general public for decades. Although extensive ecological data has been acquired, underlying molecular and cellular mechanisms remain largely unknown. In this study, to better understand mechanisms underlying gametogenesis, we isolated ovaries and testes at different developmental phases from a gonochoric coral, Euphyllia ancora, and adopted a transcriptomic approach to reveal sex- and phase-specific gene expression profiles. In particular, we explored genes associated with oocyte development and maturation, spermiogenesis, sperm motility / capacitation, and fertilization. Results 1.6 billion raw reads were obtained from 24 gonadal samples. De novo assembly of trimmed reads, and elimination of contigs derived from symbiotic dinoflagellates (Symbiodiniaceae) and other organisms yielded a reference E. ancora gonadal transcriptome of 35,802 contigs. Analysis of 4 developmental phases identified 2023 genes that were differentially expressed during oogenesis and 678 during spermatogenesis. In premature/mature ovaries, 631 genes were specifically upregulated, with 538 in mature testes. Upregulated genes included those involved in gametogenesis, gamete maturation, sperm motility / capacitation, and fertilization in other metazoans, including humans. Meanwhile, a large number of genes without homology to sequences in the SWISS-PROT database were also observed among upregulated genes in premature / mature ovaries and mature testes. Conclusions Our findings show that scleractinian gametogenesis shares many molecular characteristics with that of other metazoans, but it also possesses unique characteristics developed during cnidarian and/or scleractinian evolution. To the best of our knowledge, this study is the first to create a gonadal transcriptome assembly from any scleractinian. This study and associated datasets provide a foundation for future studies regarding gametogenesis and differences between male and female colonies from molecular and cellular perspectives. Furthermore, our transcriptome assembly will be a useful reference for future development of sex-specific and/or stage-specific germ cell markers that can be used in coral aquaculture and ecological studies.

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