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

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.

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

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.

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

PeerJ ◽  
2017 ◽  
Vol 5 ◽  
pp. e3422 ◽  
Author(s):  
Ying Wang ◽  
Kun Liu ◽  
De Bi ◽  
Shoubiao Zhou ◽  
Jianwen Shao
Keyword(s):  
Functional Annotation ◽  
De Novo ◽  
Gene Annotation ◽  
Sequence Similarity ◽  
Sequence Information ◽  
Sequencing Data ◽  
Protein Database ◽  
Illumina Hiseq ◽  
Significant Similarity ◽  
Assembly Technology

Background Desiccation-tolerant (DT) plants can recover full metabolic competence upon rehydration after losing most of their cellular water (>95%) for extended periods of time. Functional genomic approaches such as transcriptome sequencing can help us understand how DT plants survive and respond to dehydration, which has great significance for plant biology and improving the drought tolerance of crops. Boea clarkeana Hemsl. (Gesneriaceae) is a DT dicotyledonous herb. Its genomic sequences characteristics remain unknown. Based on transcriptomic analyses, polymorphic EST-SSR (simple sequence repeats in expressed sequence tags) molecular primers can be designed, which will greatly facilitate further investigations of the population genetics and demographic histories of DT plants. Methods In the present study, we used the platform Illumina HiSeq™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, the EST-SSR markers were developed, and the functional annotation of ESTs containing polymorphic SSRs were obtained through BLASTX. Results A total of 91,449 unigenes were generated from the leaf cDNA library of B. clarkeana. 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 the known proteins of 463 popular model species in the Nr database, and 59,962 unigenes and 32,336 unigenes were assigned to Gene Ontology (GO) classifications and Cluster of Orthologous Groups (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 stable, highly polymorphic loci were selected for polymorphism screening. There was a total of 65 alleles, with 2–6 alleles at each locus. Primarily due to the unique biological characteristics of plants, the HE (0–0.196), HO (0.082–0.14) and PIC (0–0.155) per locus were very low. The functional annotation distribution centered on ESTs containing di- and tri-nucleotide SSRs, and the ESTs containing primers BC2, BC4 and BC12 were annotated to vegetative dehydration/desiccation pathways. Discussion This work is the first genetic study of B. clarkeana as a new plant resource of DT genes. A substantial number of transcriptome sequences were generated in this study. These sequences are valuable resources for gene annotation and discovery as well as molecular marker development. These sequences could also provide a valuable basis for future molecular studies of B. clarkeana.

scholarly journals Transcriptome analysis of colored calla lily (Zantedeschia rehmanniiEngl.) by Illumina sequencing:de novoassembly, annotation and EST-SSR marker development

PeerJ ◽  
2016 ◽  
Vol 4 ◽  
pp. e2378 ◽  
Author(s):  
Zunzheng Wei ◽  
Zhenzhen Sun ◽  
Binbin Cui ◽  
Qixiang Zhang ◽  
Min Xiong ◽  
...  
Keyword(s):  
Molecular Markers ◽  
De Novo ◽  
Average Length ◽  
Orthologous Group ◽  
Sequence Information ◽  
Nucleotide Polymorphisms ◽  
Protein Database ◽  
Illumina Hiseq ◽  
Significant Similarity ◽  
Calla Lily

Colored calla lily is the short name for the species or hybrids in sectionAestivaeof genusZantedeschia. It is currently one of the most popular flower plants in the world due to its beautiful flower spathe and long postharvest life. However, little genomic information and few molecular markers are available for its genetic improvement. Here,de novotranscriptome sequencing was performed to produce large transcript sequences forZ. rehmanniicv. ‘Rehmannii’ using an Illumina HiSeq 2000 instrument. More than 59.9 million cDNA sequence reads were obtained and assembled into 39,298 unigenes with an average length of 1,038 bp. Among these, 21,077 unigenes showed significant similarity to protein sequences in the non-redundant protein database (Nr) and in the Swiss-Prot, Gene Ontology (GO), Cluster of Orthologous Group (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. Moreover, a total of 117 unique transcripts were then defined that might regulate the flower spathe development of colored calla lily. Additionally, 9,933 simple sequence repeats (SSRs) and 7,162 single nucleotide polymorphisms (SNPs) were identified as putative molecular markers. High-quality primers for 200 SSR loci were designed and selected, of which 58 amplified reproducible amplicons were polymorphic among 21 accessions of colored calla lily. The sequence information and molecular markers in the present study will provide valuable resources for genetic diversity analysis, germplasm characterization and marker-assisted selection in the genusZantedeschia.

scholarly journals TrancriptomeReconstructoR, A Data-Driven Annotation of Complex Transcriptomes

2020 ◽  
Author(s):  
Maxim Ivanov ◽  
Albin Sandelin ◽  
Sebastian Marquardt
Keyword(s):  
De Novo ◽  
Gene Annotation ◽  
R Package ◽  
Sequence Information ◽  
Rna Seq ◽  
Sequencing Data ◽  
Gene Model ◽  
Preparation Methods ◽  
Downstream Analysis

Abstract Background: The quality of gene annotation determines the interpretation of results obtained in transcriptomic studies. The growing number of genome sequence information calls for experimental and computational pipelines for de novo transcriptome annotation. Ideally, gene and transcript models should be called from a limited set of key experimental data. Results: We developed TranscriptomeReconstructoR, an R package which implements a pipeline for automated transcriptome annotation. It relies on integrating features from independent and complementary datasets: i) full-length RNA-seq for detection of splicing patterns and ii) high-throughput 5' and 3' tag sequencing data for accurate definition of gene borders. The pipeline can also take a nascent RNA-seq dataset to supplement the called gene model with transient transcripts.We reconstructed de novo the transcriptional landscape of wild type Arabidopsis thaliana seedlings as a proof-of-principle. A comparison to the existing transcriptome annotations revealed that our gene model is more accurate and comprehensive than the two most commonly used community gene models, TAIR10 and Araport11. In particular, we identify thousands of transient transcripts missing from the existing annotations. Our new annotation promises to improve the quality of A.thaliana genome research.Conclusions: Our proof-of-concept data suggest a cost-efficient strategy for rapid and accurate annotation of complex eukaryotic transcriptomes. We combine the choice of library preparation methods and sequencing platforms with the dedicated computational pipeline implemented in the TranscriptomeReconstructoR package. The pipeline only requires prior knowledge on the reference genomic DNA sequence, but not the transcriptome. The package seamlessly integrates with Bioconductor packages for downstream analysis.

scholarly journals TrancriptomeReconstructoR: data-driven annotation of complex transcriptomes

2020 ◽  
Author(s):  
Maxim Ivanov ◽  
Albin Sandelin ◽  
Sebastian Marquardt
Keyword(s):  
De Novo ◽  
Gene Annotation ◽  
R Package ◽  
Sequence Information ◽  
Rna Seq ◽  
Sequencing Data ◽  
Gene Model ◽  
Preparation Methods ◽  
Downstream Analysis

AbstractBackgroundThe quality of gene annotation determines the interpretation of results obtained in transcriptomic studies. The growing number of genome sequence information calls for experimental and computational pipelines for de novo transcriptome annotation. Ideally, gene and transcript models should be called from a limited set of key experimental data.ResultsWe developed TranscriptomeReconstructoR, an R package which implements a pipeline for automated transcriptome annotation. It relies on integrating features from independent and complementary datasets: i) full-length RNA-seq for detection of splicing patterns and ii) high-throughput 5’ and 3’ tag sequencing data for accurate definition of gene borders. The pipeline can also take a nascent RNA-seq dataset to supplement the called gene model with transient transcripts.We reconstructed de novo the transcriptional landscape of wild type Arabidopsis thaliana seedlings as a proof-of-principle. A comparison to the existing transcriptome annotations revealed that our gene model is more accurate and comprehensive than the two most commonly used community gene models, TAIR10 and Araport11. In particular, we identify thousands of transient transcripts missing from the existing annotations. Our new annotation promises to improve the quality of A.thaliana genome research.ConclusionsOur proof-of-concept data suggest a cost-efficient strategy for rapid and accurate annotation of complex eukaryotic transcriptomes. We combine the choice of library preparation methods and sequencing platforms with the dedicated computational pipeline implemented in the TranscriptomeReconstructoR package. The pipeline only requires prior knowledge on the reference genomic DNA sequence, but not the transcriptome. The package seamlessly integrates with Bioconductor packages for downstream analysis.

scholarly journals TrancriptomeReconstructoR: data-driven annotation of complex transcriptomes

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Maxim Ivanov ◽  
Albin Sandelin ◽  
Sebastian Marquardt
Keyword(s):  
De Novo ◽  
Gene Annotation ◽  
R Package ◽  
Sequence Information ◽  
Rna Seq ◽  
Sequencing Data ◽  
Gene Model ◽  
Preparation Methods ◽  
Downstream Analysis

Abstract Background The quality of gene annotation determines the interpretation of results obtained in transcriptomic studies. The growing number of genome sequence information calls for experimental and computational pipelines for de novo transcriptome annotation. Ideally, gene and transcript models should be called from a limited set of key experimental data. Results We developed TranscriptomeReconstructoR, an R package which implements a pipeline for automated transcriptome annotation. It relies on integrating features from independent and complementary datasets: (i) full-length RNA-seq for detection of splicing patterns and (ii) high-throughput 5′ and 3′ tag sequencing data for accurate definition of gene borders. The pipeline can also take a nascent RNA-seq dataset to supplement the called gene model with transient transcripts. We reconstructed de novo the transcriptional landscape of wild type Arabidopsis thaliana seedlings and Saccharomyces cerevisiae cells as a proof-of-principle. A comparison to the existing transcriptome annotations revealed that our gene model is more accurate and comprehensive than the most commonly used community gene models, TAIR10 and Araport11 for A.thaliana and SacCer3 for S.cerevisiae. In particular, we identify multiple transient transcripts missing from the existing annotations. Our new annotations promise to improve the quality of A.thaliana and S.cerevisiae genome research. Conclusions Our proof-of-concept data suggest a cost-efficient strategy for rapid and accurate annotation of complex eukaryotic transcriptomes. We combine the choice of library preparation methods and sequencing platforms with the dedicated computational pipeline implemented in the TranscriptomeReconstructoR package. The pipeline only requires prior knowledge on the reference genomic DNA sequence, but not the transcriptome. The package seamlessly integrates with Bioconductor packages for downstream analysis.

IsoDetect: Detection of splice isoforms from third generation long reads based on short feature sequences

2020 ◽  
Vol 15 ◽  
Author(s):  
Hongdong Li ◽  
Wenjing Zhang ◽  
Yuwen Luo ◽  
Jianxin Wang
Keyword(s):  
Sequence Similarity ◽  
Detection Methods ◽  
Sequence Information ◽  
Third Generation ◽  
Sequencing Data ◽  
Splice Isoforms ◽  
Third Generation Sequencing ◽  
Long Reads ◽  
Feature Sequence ◽  
Generation Sequencing

Aims: Accurately detect isoforms from third generation sequencing data. Background: Transcriptome annotation is the basis for the analysis of gene expression and regulation. The transcriptome annotation of many organisms such as humans is far from incomplete, due partly to the challenge in the identification of isoforms that are produced from the same gene through alternative splicing. Third generation sequencing (TGS) reads provide unprecedented opportunity for detecting isoforms due to their long length that exceeds the length of most isoforms. One limitation of current TGS reads-based isoform detection methods is that they are exclusively based on sequence reads, without incorporating the sequence information of known isoforms. Objective: Develop an efficient method for isoform detection. Method: Based on annotated isoforms, we propose a splice isoform detection method called IsoDetect. First, the sequence at exon-exon junction is extracted from annotated isoforms as the “short feature sequence”, which is used to distinguish different splice isoforms. Second, we aligned these feature sequences to long reads and divided long reads into groups that contain the same set of feature sequences, thereby avoiding the pair-wise comparison among the large number of long reads. Third, clustering and consensus generation are carried out based on sequence similarity. For the long reads that do not contain any short feature sequence, clustering analysis based on sequence similarity is performed to identify isoforms. Result: Tested on two datasets from Calypte Anna and Zebra Finch, IsoDetect showed higher speed and compelling accuracy compared with four existing methods. Conclusion: IsoDetect is a promising method for isoform detection. Other: This paper was accepted by the CBC2019 conference.

scholarly journals De Novo Transcriptomic Resources in the Brain of Vespa velutina for Invasion Control

Insects ◽  
2020 ◽  
Vol 11 (2) ◽  
pp. 101
Author(s):  
Miao Wang ◽  
Hanyu Li ◽  
Huoqing Zheng ◽  
Liuwei Zhao ◽  
Xiaofeng Xue ◽  
...  
Keyword(s):  
Large Scale ◽  
Target Genes ◽  
De Novo ◽  
Transcriptome Profiling ◽  
Molecular Study ◽  
Snp Markers ◽  
Nucleotide Polymorphisms ◽  
Illumina Hiseq ◽  
Vespa Velutina ◽  
Genes Encoding

The invasion of Vespa velutina presents a great threat to the agriculture economy, the ecological environment, and human health. An effective strategy for this hornet control is urgently required, but the limited genome information of Vespa velutina restricts the application of molecular-genomic tools for targeted hornet management. Therefore, we conducted large-scale transcriptome profiling of the hornet brain to obtain functional target genes and molecular markers. Using an Illumina HiSeq platform, more than 41 million clean reads were obtained and de novo assembled into 182,087 meaningful unigenes. A total of 56,400 unigenes were annotated against publicly available protein sequence databases and a set of reliable Simple Sequence Repeats (SSRs) and Single Nucleotide Polymorphisms (SNP) markers were developed. The homologous genes encoding crucial behavior regulation factors, odorant binding proteins (OBPs), and vitellogenin, were also identified from highly expressed transcripts. This study provides abundant molecular targets and markers for invasive hornet control and further promotes the genetic and molecular study of Vespa velutina.

scholarly journals Overview of organohalide-respiring bacteria and a proposal for a classification system for reductive dehalogenases

2013 ◽  
Vol 368 (1616) ◽  
pp. 20120322 ◽  
Author(s):  
Laura A. Hug ◽  
Farai Maphosa ◽  
David Leys ◽  
Frank E. Löffler ◽  
Hauke Smidt ◽  
...  
Keyword(s):  
Classification System ◽  
Sequence Similarity ◽  
Halogenated Hydrocarbons ◽  
Sequence Information ◽  
Sequencing Data ◽  
Functional Prediction ◽  
Groundwater Contaminants ◽  
Reductive Dehalogenase ◽  
Bacterial Phyla ◽  
Reductive Dehalogenase Genes

Organohalide respiration is an anaerobic bacterial respiratory process that uses halogenated hydrocarbons as terminal electron acceptors during electron transport-based energy conservation. This dechlorination process has triggered considerable interest for detoxification of anthropogenic groundwater contaminants. Organohalide-respiring bacteria have been identified from multiple bacterial phyla, and can be categorized as obligate and non-obligate organohalide respirers. The majority of the currently known organohalide-respiring bacteria carry multiple reductive dehalogenase genes. Analysis of a curated set of reductive dehalogenases reveals that sequence similarity and substrate specificity are generally not correlated, making functional prediction from sequence information difficult. In this article, an orthologue-based classification system for the reductive dehalogenases is proposed to aid integration of new sequencing data and to unify terminology.

scholarly journals Multi-tissue transcriptome analysis using hybrid-seq revealed potential genes and biological pathways associated with azadirachtin A biosynthesis in neem (Azadirachtin indica)

2020 ◽  
Author(s):  
Huiyan Wang ◽  
Ning Wang ◽  
Yixin Huo
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Biosynthetic Pathway ◽  
De Novo ◽  
Analysis Data ◽  
Sequencing Data ◽  
Neem Tree ◽  
Illumina Hiseq ◽  
Oxidosqualene Cyclase ◽  
Azadirachtin A

Abstract Background: Azadirachtin A is a triterpenoid from neem tree exhibiting excellent activities against over 600 insect species in agriculture. The manufacture of azadirachtin A depends on extraction from neem tissues, which is not ecofriendly and sustainable. The low yield and discontinuous supply impeded the further application. The biosynthetic pathway of azadirachtin A is still well-known.Results: We attempted to explore azadirachtin A biosynthetic pathway and identified key involved genes by analyzing transcriptome data of five neem tissues through hybrid-seq (Illumina HiSeq and Pacific Biosciences Single Molecule Real Time (PacBio SMRT)) technology. A total 219 and 397 up-regulated differentially expressed genes (DEGs) in leaf and fruit tissues than other tissues (root, stem and flower) were isolated. After phylogenetic analysis and domain prediction, 22 candidates encoding 2,3-oxidosqualene cyclase (OSC), alcohol dehydrogenase (ADH), cytochrome P450 (CYP450), acyltransferase (ACT) and esterase (EST) proposed to be involved in azadirachtin A biosynthesis were finally selected. De novo assembled sequences were verified by Quantitative Real-Time PCR (qRT-PCR) analysis.Conclusions: By integrating and analysis data from Illumina HiSeq and PacBio SMRT platform, 22 DEGs were finally selected as candidates involved in azadirachtin A biosynthesis. The obtained reliable and accurate sequencing data provided important novel information for understanding neem genome. Our data shed new light on the understanding of other triterpenoids biosynthesis in neem trees and provide reference for exploring other valuable natural product biosynthesis in plants.

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