Analyses of MYMIV-induced transcriptome in Vigna mungo as revealed by next generation sequencing.
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ABSTRACT:
SUBMITTER: Ganguli S
PROVIDER: S-EPMC4778624 | biostudies-literature | 2016 Mar
REPOSITORIES: biostudies-literature
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Project description:Mungbean Yellow Mosaic India Virus (MYMIV)-infection creates major hindrance in V. mungo cultivation and poses significant threat to other grain legume production. Symptoms associated include severe patho-physiological alterations characterized by chlorotic foliar lesion accompanied by reduced growth. However, dissection of the host's defense machinery remains a tough challenge due to limited of host's genomic resources. A comparative RNA-Seq transcriptomes of resistant (VM84) and susceptible (T9) plants was carried out to identify genes potentially involved in V. mungo resistance against MYMIV. Distinct gene expression landscapes were observed in VM84 and T9 with 2158 and 1679 differentially expressed genes (DEGs), respectively. Transcriptomic responses in VM84 reflect a prompt and intense immune reaction demonstrating an efficient pathogen surveillance leading to activation of basal and induced immune responses. Functional analysis of the altered DEGs identified multiple regulatory pathways to be activated or repressed over time. Up-regulation of DEGs including NB-LRR, WRKY33, ankyrin, argonaute and NAC transcription factor revealed an insight on their potential roles in MYMIV-resistance; and qPCR validation shows a propensity of their accumulation in VM84. Analyses of the current RNA-Seq dataset contribute immensely to decipher molecular responses that underlie MYMIV-resistance and will aid in the improvement strategy of V. mungo and other legumes through comparative functional genomics.
| S-EPMC6586629 | biostudies-literature
Project description:Vigna mungo is cultivated in approximately 5 million hectares worldwide. The chloroplast genome of this species has not been previously reported. In this study, we sequenced the genome and transcriptome of the V. mungo chloroplast. We identified many positively selected genes in the photosynthetic pathway (e.g., rbcL, ndhF, and atpF) and RNA polymerase genes (e.g., rpoC2) from the comparison of the chloroplast genome of V. mungo, temperate legume species, and tropical legume species. Our transcriptome data from PacBio isoform sequencing showed that the 51-kb DNA inversion could affect the transcriptional regulation of accD polycistronic. Using Illumina deep RNA sequencing, we found RNA editing of clpP in the leaf, shoot, flower, fruit, and root tissues of V. mungo. We also found three G-to-A RNA editing events that change guanine to adenine in the transcripts transcribed from the adenine-rich regions of the ycf4 gene. The edited guanine bases were found particularly in the chloroplast genome of the Vigna species. These G-to-A RNA editing events were likely to provide a mechanism for correcting DNA base mutations. The V. mungo chloroplast genome sequence and the analysis results obtained in this study can apply to phylogenetic studies and chloroplast genome engineering.
| S-EPMC7570002 | biostudies-literature
Project description:Initial phases of the MYMIV-Vigna mungo interaction is crucial in determining the infection phenotype upon challenging with the virus. During incompatible interaction, the plant deploys multiple stratagems that include extensive transcriptional alterations defying the virulence factors of the pathogen. Such molecular events are not frequently addressed by genomic tools. In order to obtain a critical insight to unravel how V. mungo respond to Mungbean yellow mosaic India virus (MYMIV), we have employed the PCR based suppression subtractive hybridization technique to identify genes that exhibit altered expressions. Dynamics of 345 candidate genes are illustrated that differentially expressed either in compatible or incompatible reactions and their possible biological and cellular functions are predicted. The MYMIV-induced physiological aspects of the resistant host include reactive oxygen species generation, induction of Ca2+ mediated signaling, enhanced expression of transcripts involved in phenylpropanoid and ubiquitin-proteasomal pathways; all these together confer resistance against the invader. Elicitation of genes implicated in salicylic acid (SA) pathway suggests that immune response is under the regulation of SA signaling. A significant fraction of modulated transcripts are of unknown function indicating participation of novel candidate genes in restricting this viral pathogen. Susceptibility on the other hand, as exhibited by V. mungo Cv. T9 is perhaps due to the poor execution of these transcript modulation exhibiting remarkable repression of photosynthesis related genes resulting in chlorosis of leaves followed by penalty in crop yield. Thus, the present findings revealed an insight on the molecular warfare during host-virus interaction suggesting plausible signaling mechanisms and key biochemical pathways overriding MYMIV invasion in resistant genotype of V. mungo. In addition to inflate the existing knowledge base, the genomic resources identified in this orphan crop would be useful for integrating MYMIV-tolerance trait in susceptible cultivars of V. mungo.
| S-EPMC4401676 | biostudies-literature
Project description:NGPS is a method for de-novo, full-length protein sequencing in high throughput. The method is based on cleavage of the protein at semi-random sites by microwave-assisted acid hydrolysis (MAAH), enrichment of LC-MS/MS amenable peptides from the hydrolysate by solid-phase-extraction, LC-MS/MS analysis, de-novo long peptide tag sequencing of resulting peptides and assembly of peptide tags into consensus contigs.
2017-04-03 | PXD003804 | Pride
Project description:BACKGROUND:Next-Generation Sequencing (NGS) technology has exceptionally increased the ability to sequence DNA in a massively parallel and cost-effective manner. Nevertheless, NGS data analysis requires bioinformatics skills and computational resources well beyond the possibilities of many "wet biology" laboratories. Moreover, most of projects only require few sequencing cycles and standard tools or workflows to carry out suitable analyses for the identification and annotation of genes, transcripts and splice variants found in the biological samples under investigation. These projects can take benefits from the availability of easy to use systems to automatically analyse sequences and to mine data without the preventive need of strong bioinformatics background and hardware infrastructure. RESULTS:To address this issue we developed an automatic system targeted to the analysis of NGS data obtained from large-scale transcriptome studies. This system, we named NGS-Trex (NGS Transcriptome profile explorer) is available through a simple web interface http://www.ngs-trex.org and allows the user to upload raw sequences and easily obtain an accurate characterization of the transcriptome profile after the setting of few parameters required to tune the analysis procedure. The system is also able to assess differential expression at both gene and transcript level (i.e. splicing isoforms) by comparing the expression profile of different samples.By using simple query forms the user can obtain list of genes, transcripts, splice sites ranked and filtered according to several criteria. Data can be viewed as tables, text files or through a simple genome browser which helps the visual inspection of the data. CONCLUSIONS:NGS-Trex is a simple tool for RNA-Seq data analysis mainly targeted to "wet biology" researchers with limited bioinformatics skills. It offers simple data mining tools to explore transcriptome profiles of samples investigated taking advantage of NGS technologies.
| S-EPMC3633008 | biostudies-literature
Project description:Whole genome resequencing of blackgram (Vigna mungo)
| PRJNA858573 | ENA
Project description:BackgroundMitochondrial genomes comprise a small but critical component of the total DNA in eukaryotic organisms. They encode several key proteins for the cell's major energy producing apparatus, the mitochondrial respiratory chain. Additionally, their nucleotide and amino acid sequences are of great utility as markers for systematics, molecular ecology and forensics. Their characterization through nucleotide sequencing is a fundamental starting point in mitogenomics. Methods to amplify complete mitochondrial genomes rapidly and efficiently from microgram quantities of tissue of single individuals are, however, not always available. Here we validate two approaches, which combine long-PCR with Roche 454 pyrosequencing technology, to obtain two complete mitochondrial genomes from individual amphibian species.ResultsWe obtained two new xenopus frogs (Xenopus borealis and X. victorianus) complete mitochondrial genome sequences by means of long-PCR followed by 454 of individual genomes (approach 1) or of multiple pooled genomes (approach 2), the mean depth of coverage per nucleotide was 9823 and 186, respectively. We also characterised and compared the new mitogenomes against their sister taxa; X. laevis and Silurana tropicalis, two of the most intensely studied amphibians. Our results demonstrate how our approaches can be used to obtain complete amphibian mitogenomes with depths of coverage that far surpass traditional primer-walking strategies, at either the same cost or less. Our results also demonstrate: that the size, gene content and order are the same among xenopus mitogenomes and that S. tropicalis form a separate clade to the other xenopus, among which X. laevis and X. victorianus were most closely related. Nucleotide and amino acid diversity was found to vary across the xenopus mitogenomes, with the greatest diversity observed in the Complex 1 gene nad4l and the least diversity observed in Complex 4 genes (cox1-3). All protein-coding genes were shown to be under strong negative (purifying selection), with genes under the strongest pressure (Complex 4) also being the most highly expressed, highlighting their potentially crucial functions in the mitochondrial respiratory chain.ConclusionsNext generation sequencing of long-PCR amplicons using single taxon or multi-taxon approaches enabled two new species of Xenopus mtDNA to be fully characterized. We anticipate our complete mitochondrial genome amplification methods to be applicable to other amphibians, helpful for identifying the most appropriate markers for differentiating species, populations and resolving phylogenies, a pressing need since amphibians are undergoing drastic global decline. Our mtDNAs also provide templates for conserved primer design and the assembly of RNA and DNA reads following high throughput "omic" techniques such as RNA- and ChIP-seq. These could help us better understand how processes such mitochondrial replication and gene expression influence xenopus growth and development, as well as how they evolved and are regulated.
| S-EPMC3546946 | biostudies-literature
Project description:Black gram (Vigna mungo) seeds are a rich source of digestible proteins, however, during storage these seeds are severely damaged by bruchids (Callosobruchus spp.), reducing seed quality and yield losses. Most of the cultivated genotypes of black gram are susceptible to bruchids, however, few tolerant genotypes have also been identified but the mechanism of tolerance is poorly understood. We employed Suppression Subtractive Hybridization (SSH) to identify specifically, but rarely expressed bruchid egg induced genes in black gram. In this study, Suppression Subtractive Hybridization (SSH) library was constructed to study the genes involved in defense response in black gram against bruchid infestation. An EST library of 277 clones was obtained for further analyses. Based on CAP3 assembly, 134 unigenes were computationally annotated using Blast2GOPRO software. In all, 20 defense related genes were subject to quantitative PCR analysis (qPCR) out of which 12 genes showed up-regulation in developing seeds of the pods oviposited by bruchids. Few major defense genes like defensin, pathogenesis related protein (PR), lipoxygenase (LOX) showed high expression levels in the oviposited population when compared with the non-oviposited plants. This is the first report on defense related gene transcript dynamics during the bruchid-black gram interaction using SSH library. This library would be useful to clone defense related gene(s) such as defensin as represented in our library for crop improvement.
| S-EPMC5407641 | biostudies-literature
Project description:BACKGROUND: We have developed a simulation approach to help determine the optimal mixture of sequencing methods for most complete and cost effective transcriptome sequencing. We compared simulation results for traditional capillary sequencing with "Next Generation" (NG) ultra high-throughput technologies. The simulation model was parameterized using mappings of 130,000 cDNA sequence reads to the Arabidopsis genome (NCBI Accession SRA008180.19). We also generated 454-GS20 sequences and de novo assemblies for the basal eudicot California poppy (Eschscholzia californica) and the magnoliid avocado (Persea americana) using a variety of methods for cDNA synthesis. RESULTS: The Arabidopsis reads tagged more than 15,000 genes, including new splice variants and extended UTR regions. Of the total 134,791 reads (13.8 MB), 119,518 (88.7%) mapped exactly to known exons, while 1,117 (0.8%) mapped to introns, 11,524 (8.6%) spanned annotated intron/exon boundaries, and 3,066 (2.3%) extended beyond the end of annotated UTRs. Sequence-based inference of relative gene expression levels correlated significantly with microarray data. As expected, NG sequencing of normalized libraries tagged more genes than non-normalized libraries, although non-normalized libraries yielded more full-length cDNA sequences. The Arabidopsis data were used to simulate additional rounds of NG and traditional EST sequencing, and various combinations of each. Our simulations suggest a combination of FLX and Solexa sequencing for optimal transcriptome coverage at modest cost. We have also developed ESTcalc http://fgp.huck.psu.edu/NG_Sims/ngsim.pl, an online webtool, which allows users to explore the results of this study by specifying individualized costs and sequencing characteristics. CONCLUSION: NG sequencing technologies are a highly flexible set of platforms that can be scaled to suit different project goals. In terms of sequence coverage alone, the NG sequencing is a dramatic advance over capillary-based sequencing, but NG sequencing also presents significant challenges in assembly and sequence accuracy due to short read lengths, method-specific sequencing errors, and the absence of physical clones. These problems may be overcome by hybrid sequencing strategies using a mixture of sequencing methodologies, by new assemblers, and by sequencing more deeply. Sequencing and microarray outcomes from multiple experiments suggest that our simulator will be useful for guiding NG transcriptome sequencing projects in a wide range of organisms.
| S-EPMC2907694 | biostudies-literature