Relevant bibliographies by topics / Ribo-depletion

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Academic literature on the topic 'Ribo-depletion'

Author: Grafiati
Published: 4 June 2021
Last updated: 12 February 2022

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Journal articles on the topic "Ribo-depletion"

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Alkan, Ferhat, Joana Silva, Eric Pintó Barberà, and William J. Faller. "Ribo-ODDR: oligo design pipeline for experiment-specific rRNA depletion in Ribo-seq." Bioinformatics 37, no. 17 (2021): 2659–67. http://dx.doi.org/10.1093/bioinformatics/btab171.

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Abstract Motivation Ribosome Profiling (Ribo-seq) has revolutionized the study of RNA translation by providing information on ribosome positions across all translated RNAs with nucleotide-resolution. Yet several technical limitations restrict the sequencing depth of such experiments, the most common of which is the overabundance of rRNA fragments. Various strategies can be employed to tackle this issue, including the use of commercial rRNA depletion kits. However, as they are designed for more standardized RNAseq experiments, they may perform suboptimally in Ribo-seq. In order to overcome this, it is possible to use custom biotinylated oligos complementary to the most abundant rRNA fragments, however currently no computational framework exists to aid the design of optimal oligos. Results Here, we first show that a major confounding issue is that the rRNA fragments generated via Ribo-seq vary significantly with differing experimental conditions, suggesting that a ‘one-size-fits-all’ approach may be inefficient. Therefore we developed Ribo-ODDR, an oligo design pipeline integrated with a user-friendly interface that assists in oligo selection for efficient experiment-specific rRNA depletion. Ribo-ODDR uses preliminary data to identify the most abundant rRNA fragments, and calculates the rRNA depletion efficiency of potential oligos. We experimentally show that Ribo-ODDR designed oligos outperform commercially available kits and lead to a significant increase in rRNA depletion in Ribo-seq. Availability and implementation Ribo-ODDR is freely accessible at https://github.com/fallerlab/Ribo-ODDR. Supplementary information Supplementary data are available at Bioinformatics online.
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Guo, Yan, Jie Wu, Shilin Zhao, et al. "RNA Sequencing of Formalin-Fixed, Paraffin-Embedded Specimens for Gene Expression Quantification and Data Mining." International Journal of Genomics 2016 (2016): 1–10. http://dx.doi.org/10.1155/2016/9837310.

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Background. Proper rRNA depletion is crucial for the successful utilization of FFPE specimens when studying gene expression. We performed a study to evaluate two major rRNA depletion methods: Ribo-Zero and RNase H. RNAs extracted from 4 samples were treated with the two rRNA depletion methods in duplicate and sequenced (N=16). We evaluated their reducibility, ability to detect RNA, and ability to molecularly subtype these triple negative breast cancer specimens.Results. Both rRNA depletion methods produced consistent data between the technical replicates. We found that the RNase H method produced higher quality RNAseq data as compared to the Ribo-Zero method. In addition, we evaluated the RNAseq data generated from the FFPE tissue samples for noncoding RNA, including lncRNA, enhancer/super enhancer RNA, and single nucleotide variation (SNV). We found that the RNase H is more suitable for detecting high-quality, noncoding RNAs as compared to the Ribo-Zero and provided more consistent molecular subtype identification between replicates. Unfortunately, neither method produced reliable SNV data.Conclusions. In conclusion, for FFPE specimens, the RNase H rRNA depletion method performed better than the Ribo-Zero. Neither method generates data sufficient for SNV detection.
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Thompson, Mary Kay, Maria Kiourlappou, and Ilan Davis. "Ribo-Pop: simple, cost-effective, and widely applicable ribosomal RNA depletion." RNA 26, no. 11 (2020): 1731–42. http://dx.doi.org/10.1261/rna.076562.120.

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Glaub, Alina, Christopher Huptas, Klaus Neuhaus, and Zachary Ardern. "Recommendations for bacterial ribosome profiling experiments based on bioinformatic evaluation of published data." Journal of Biological Chemistry 295, no. 27 (2020): 8999–9011. http://dx.doi.org/10.1074/jbc.ra119.012161.

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Ribosome profiling (RIBO-Seq) has improved our understanding of bacterial translation, including finding many unannotated genes. However, protocols for RIBO-Seq and corresponding data analysis are not yet standardized. Here, we analyzed 48 RIBO-Seq samples from nine studies of Escherichia coli K12 grown in lysogeny broth medium and particularly focused on the size-selection step. We show that for conventional expression analysis, a size range between 22 and 30 nucleotides is sufficient to obtain protein-coding fragments, which has the advantage of removing many unwanted rRNA and tRNA reads. More specific analyses may require longer reads and a corresponding improvement in rRNA/tRNA depletion. There is no consensus about the appropriate sequencing depth for RIBO-Seq experiments in prokaryotes, and studies vary significantly in total read number. Our analysis suggests that 20 million reads that are not mapping to rRNA/tRNA are required for global detection of translated annotated genes. We also highlight the influence of drug-induced ribosome stalling, which causes bias at translation start sites. The resulting accumulation of reads at the start site may be especially useful for detecting weakly expressed genes. As different methods suit different questions, it may not be possible to produce a “one-size-fits-all” ribosome profiling data set. Therefore, experiments should be carefully designed in light of the scientific questions of interest. We propose some basic characteristics that should be reported with any new RIBO-Seq data sets. Careful attention to the factors discussed should improve prokaryotic gene detection and the comparability of ribosome profiling data sets.
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Telzrow, Calla L., Paul J. Zwack, Shannon Esher Righi, et al. "Comparative analysis of RNA enrichment methods for preparation of Cryptococcus neoformans RNA sequencing libraries." G3 Genes|Genomes|Genetics, August 26, 2021. http://dx.doi.org/10.1093/g3journal/jkab301.

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Abstract RNA sequencing (RNA-Seq) experiments focused on gene expression involve removal of ribosomal RNA (rRNA) because it is the major RNA constituent of cells. This process, called RNA enrichment, is done primarily to reduce cost: without rRNA removal, deeper sequencing must be performed to compensate for the sequencing reads wasted on rRNA. The ideal RNA enrichment method removes all rRNA without affecting other RNA in the sample. We tested the performance of three RNA enrichment methods on RNA isolated from Cryptococcus neoformans, a fungal pathogen of humans. We find that the RNase H depletion method is more efficient in depleting rRNA and more specific in recapitulating non-rRNA levels present in unenriched controls than the commonly-used Poly(A) isolation method. The RNase H depletion method is also more effective than the Ribo-Zero depletion method as measured by rRNA depletion efficiency and recapitulation of protein-coding RNA levels present in unenriched controls, while the Ribo-Zero depletion method more closely recapitulates annotated non-coding RNA (ncRNA) levels. Finally, we leverage these data to accurately map the C. neoformans mitochondrial rRNA genes, and also demonstrate that RNA-Seq data generated with the RNase H and Ribo-Zero depletion methods can be used to explore novel C. neoformans long non-coding RNA genes.
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Engelhardt, Florian, Jürgen Tomasch, and Susanne Häussler. "Organism-specific depletion of highly abundant RNA species from bacterial total RNA." Access Microbiology 2, no. 10 (2020). http://dx.doi.org/10.1099/acmi.0.000159.

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High-throughput sequencing has become a standard tool for transcriptome analysis. The depletion of overrepresented RNA species from sequencing libraries plays a key role in establishing potent and cost-efficient RNA-seq routines. Commercially available kits are known to obtain good results for the reduction of ribosomal RNA (rRNA). However, we found that the transfer-messenger RNA (tmRNA) was frequently highly abundant in rRNA-depleted samples of Pseudomonas aeruginosa , consuming up to 25 % of the obtained reads. The tmRNA fraction was particularly high in samples taken from stationary cultures. This suggests that overrepresentation of this RNA species reduces the mRNA fraction when cells are grown under challenging conditions. Here, we present an RNase-H-based depletion protocol that targets the tmRNA in addition to ribosomal RNAs. We were able to increase the mRNA fraction to 93–99% and therefore outperform not only the commercially Ribo-off kit (Vazyme) operating by the same principle but also the formerly widely used Ribo-Zero kit (Illumina). Maximizing the read share of scientifically interesting RNA species enhances the discriminatory potential of next-generation RNA-seq experiments and, therefore, can contribute to a better understanding of the transcriptomic landscape of bacterial pathogens and their used mechanisms in host infection.
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Jang, Jin Sung, Brianna Berg, Eileen Holicky, et al. "Comparative evaluation for the globin gene depletion methods for mRNA sequencing using the whole blood-derived total RNAs." BMC Genomics 21, no. 1 (2020). http://dx.doi.org/10.1186/s12864-020-07304-4.

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Abstract Background There are challenges in generating mRNA-Seq data from whole-blood derived RNA as globin gene and rRNA are frequent contaminants. Given the abundance of erythrocytes in whole blood, globin genes comprise some 80% or more of the total RNA. Therefore, depletion of globin gene RNA and rRNA are critical steps required to have adequate coverage of reads mapping to the reference transcripts and thus reduce the total cost of sequencing. In this study, we directly compared the performance of probe hybridization (GLOBINClear Kit and Globin-Zero Gold rRNA Removal Kit) and RNAse-H enzymatic depletion (NEBNext® Globin & rRNA Depletion Kit and Ribo-Zero Plus rRNA Depletion Kit) methods from 1 μg of whole blood-derived RNA on mRNA-Seq profiling. All RNA samples were treated with DNaseI for additional cleanup before the depletion step and were processed for poly-A selection for library generation. Results Probe hybridization revealed a better overall performance than the RNAse-H enzymatic depletion method, detecting a higher number of genes and transcripts without 3′ region bias. After depletion, samples treated with probe hybridization showed globin genes at 0.5% (±0.6%) of the total mapped reads; the RNAse-H enzymatic depletion had 3.2% (±3.8%). Probe hybridization showed more junction reads and transcripts compared with RNAse-H enzymatic depletion and also had a higher correlation (R > 0.9) than RNAse-H enzymatic depletion (R > 0.85). Conclusion In this study, our results showed that 1 μg of high-quality RNA from whole blood could be routinely used for transcriptional profiling analysis studies with globin gene and rRNA depletion pre-processing. We also demonstrated that the probe hybridization depletion method is better suited to mRNA sequencing analysis with minimal effect on RNA quality during depletion procedures.
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Eshraghi, Mehdi, Pabalu P. Karunadharma, Juliana Blin, et al. "Mutant Huntingtin stalls ribosomes and represses protein synthesis in a cellular model of Huntington disease." Nature Communications 12, no. 1 (2021). http://dx.doi.org/10.1038/s41467-021-21637-y.

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AbstractThe polyglutamine expansion of huntingtin (mHTT) causes Huntington disease (HD) and neurodegeneration, but the mechanisms remain unclear. Here, we found that mHtt promotes ribosome stalling and suppresses protein synthesis in mouse HD striatal neuronal cells. Depletion of mHtt enhances protein synthesis and increases the speed of ribosomal translocation, while mHtt directly inhibits protein synthesis in vitro. Fmrp, a known regulator of ribosome stalling, is upregulated in HD, but its depletion has no discernible effect on protein synthesis or ribosome stalling in HD cells. We found interactions of ribosomal proteins and translating ribosomes with mHtt. High-resolution global ribosome footprint profiling (Ribo-Seq) and mRNA-Seq indicates a widespread shift in ribosome occupancy toward the 5′ and 3′ end and unique single-codon pauses on selected mRNA targets in HD cells, compared to controls. Thus, mHtt impedes ribosomal translocation during translation elongation, a mechanistic defect that can be exploited for HD therapeutics.
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Harrington, Christina A., Suzanne S. Fei, Jessica Minnier, et al. "RNA-Seq of human whole blood: Evaluation of globin RNA depletion on Ribo-Zero library method." Scientific Reports 10, no. 1 (2020). http://dx.doi.org/10.1038/s41598-020-62801-6.

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Newton, Yulia, Andrew J. Sedgewick, Luis Cisneros, et al. "Large scale, robust, and accurate whole transcriptome profiling from clinical formalin-fixed paraffin-embedded samples." Scientific Reports 10, no. 1 (2020). http://dx.doi.org/10.1038/s41598-020-74483-1.

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Abstract Transcriptome profiling can provide information of great value in clinical decision-making, yet RNA from readily available formalin-fixed paraffin-embedded (FFPE) tissue is often too degraded for quality sequencing. To assess the clinical utility of FFPE-derived RNA, we performed ribo-deplete RNA extractions on > 3200 FFPE slide samples; 25 of these had direct FFPE vs. fresh frozen (FF) replicates, 57 were sequenced in 2 different labs, 87 underwent multiple library analyses, and 16 had direct microdissected vs. macrodissected replicates. Poly-A versus ribo-depletion RNA extraction methods were compared using transcriptomes of TCGA cohort and 3116 FFPE samples. Compared to FF, FFPE transcripts coding for nuclear/cytoplasmic proteins involved in DNA packaging, replication, and protein synthesis were detected at lower rates and zinc finger family transcripts were of poorer quality. The greatest difference in extraction methods was in histone transcripts which typically lack poly-A tails. Encouragingly, the overall sequencing success rate was 81%. Exome coverage was highly concordant in direct FFPE and FF replicates, with 98% agreement in coding exon coverage and a median correlation of whole transcriptome profiles of 0.95. We provide strong rationale for clinical use of FFPE-derived RNA based on the robustness, reproducibility, and consistency of whole transcriptome profiling.
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Dissertations / Theses on the topic "Ribo-depletion"

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Peterson, Brittany F., and Michael E. Scharf. "Metatranscriptome analysis reveals bacterial symbiont contributions to lower termite physiology and potential immune functions." BIOMED CENTRAL LTD, 2016. http://hdl.handle.net/10150/621516.

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Background: Symbioses throughout the animal kingdom are known to extend physiological and ecological capabilities to hosts. Insect-microbe associations are extremely common and are often related to novel niche exploitation, fitness advantages, and even speciation events. These phenomena include expansions in host diet, detoxification of insecticides and toxins, and increased defense against pathogens. However, dissecting the contributions of individual groups of symbionts at the molecular level is often underexplored due to methodological and analytical limitations. Termites are one of the best studied systems for physiological collaborations between host and symbiota; however, most work in lower termites (those with bacterial and protist symbionts) focuses on the eukaryotic members of this symbiotic consortium. Here we present a metatranscriptomic analysis which provides novel insights into bacterial contributions to the holobiont of the eastern subterranean termite, Reticulitermes flavipes, in the presence and absence of a fungal pathogen. Results: Using a customized ribodepletion strategy, a metatranscriptome assembly was obtained representing the host termite as well as bacterial and protist symbiota. Sequence data provide new insights into biosynthesis, catabolism, and transport of major organic molecules and ions by the gut consortium, and corroborate previous findings suggesting that bacteria play direct roles in nitrogen fixation, amino acid biosynthesis, and lignocellulose digestion. With regard to fungal pathogen challenge, a total of 563 differentially expressed candidate host and symbiont contigs were identified (162 up-and 401 downregulated; a/FDR = 0.05) including an upregulated bacterial amidohydrolase. Conclusions: This study presents the most complete bacterial metatranscriptome from a lower termite and provides a framework on which to build a more complete model of termite-symbiont interactions including, but not limited to, digestion and pathogen defense.
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Conference papers on the topic "Ribo-depletion"

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Kumar, Ashwini, Matti Kankainen, Alun Parsons, Olli Kallioniemi, Pirkko Mattila, and Caroline Heckman. "Abstract 1517: Impact of poly-A and ribo-depletion RNA-seq library construction protocols on transcriptomic analysis of samples from patients with haematological malignancies." In Proceedings: AACR 107th Annual Meeting 2016; April 16-20, 2016; New Orleans, LA. American Association for Cancer Research, 2016. http://dx.doi.org/10.1158/1538-7445.am2016-1517.

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